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CHEMICAL ANALYSIS 



OF 



Healthy and Diseased Urine, 



QUALITATIVE AND QUANTITATIVE. 



a 



T. C. VAN NUYS, 



PROFESSOR OF CHEMISTRY, INDIANA UNIVERSITY, 



7^ 



THIRTY-NINE WOOD ENGRAVINGS 

mogp 
JAN 20 1888 

PHILADELPHIA : 

P. BLAKISTON, SON & CO. 

1012 Walnut Street. 
1888. 






Copyright, 1887, by P. Blakiston, Son & Co. 



PREFACE 



In the preparation of this Manual no labor has been spared to 
adapt it to the requirements of all persons into whose hands it 
may fall. With this object in view, simple qualitative tests are 
given in full, and the rationale of chemical processes fully 
explained, and, to meet the requirements of students somewhat 
familiar with laboratory work, nearly all the methods employed 
in quantitative estimations are given in full; especially is this 
true in processes peculiar to work in physiological chemistry. 

The plan of the part of the work devoted to the Qualitative 
Examination of the Urine is understood by examination of the 
Index of Chapters. Chapter VI is devoted to Processes of 
Examination of the Urine and Sediments, and Chapter VII is 
given to the consideration of Concretions and Stones. The pre- 
ceding chapters, I to V inclusive, are devoted to the Physical and 
Chemical Properties of Constituents of the Urine. These chap- 
ters, therefore, embrace materials with which the student should 
become quite familiar by experimentation, in order that the work 
laid out in Chapters VI and VII may not be mechanical. 

In Chapter VIII is found a brief consideration of Processes and 
Apparatus employed in Quantitative work, and also the Prepa- 
ration of Normal Solutions required. In this connection the 
student should not infer that a general work on Quantitative 
Analysis, embracing details of manipulation with description of 
apparatus, is not required. The most important facts, as regards 
laboratory work, have been incorporated, that they may be of 
some assistance to those who have no complete treatise at hand. 



IV PREFACE. 



In Chapters IX, X and XI are found Methods for Estimation 
of Quantities of Constituents of Healthy or Diseased Urine, as the 
case may be, and in Chapter XII are Methods for Estimation of 
Quantities of Albuminous Bodies, and Sugar in Urine. As an aid 
to students to gain an insight into the constitution of food and 
urine, and the relationship they sustain to each other, the four 
Tables in the Appendix are introduced. Tables 2, 3 and 4 are 
from Zuelzer, while Table 1 was arranged from data collected by 
the author. 

The Quantitative part of the work is perhaps fuller than is 
generally required, but with a moment's consideration it is under- 
stood that it was by quantitative analysis that nearly all the facts 
concerning the transformation of tissue and the elements of food 
have been brought to light; and that quantitative analysis will 
eventually be employed in many cases as an aid in diagnosis and 
in the treatment of disease, is an assured fact if the practice of 
medicine is to become to a certain extent fixed, as mathematics 
and physics. 

Presumably it would not be speculative to assume that the 
natural history of a specific disease embraces a typical composi- 
tion of the urine.' In this direction much is due to the labors of 
Zuelzer, of Germany, and Lepine, of France. By Zuelzer the 
quantity of nitrogen excreted by the kidneys is placed at 100, 
and in health the variation of the relative quantity of each 
constituent of the urine is known from the results of numerous 
estimations. This basis of measurement is satisfactory in all cases, 
as nitrogen is a constituent of all tissues, and nitrogenous pro- 
ducts are excreted by the kidneys. When an organ or particular 
tissue yields an increased quantity of products of waste these 
products are, to a great extent, found in the urine, in which case 
the relative quantities of these products are increased. Without 
some standard of measurement, the fact that a product, P 2 5 for 
example, is found in increased quantity in the urine, is of little or 



PREFACE. V 

no value ; but if the quantity of P 2 5 is much greater than in 
health, while that of nitrogen is not greatly increased, attention is 
directed to tissues containing much phosphorus, as lecithin, in 
nerve centres. From these facts it is implied that the estimation 
of the quantity of nitrogen in urine should be made, when the 
quantities of, other constituents are determined. 

Until recently this was an obstacle not easily overcome, when 
the only methods for the estimation of nitrogen leading to correct 
results were those of Dumas and Varrentrapp and Will, but now, 
as Kjeldahl's method is known to yield results as accurate and 
not requiring much time or apparatus, there is no reason why the 
employment of quantitative estimations will not lead to a knowl- 
edge of types in the constitution of the urine peculiar to different 
diseases. It was with this view that the quantitative part of the 
work was made somewhat lengthy, and that the methods em- 
ployed are the most exact. 

Van Nuys. 

Indiana University Chemical Laboratory, 
Bloomington, Ind., December, 1887. 



CONTENTS 



CHAPTER I. 



Properties of Normal Urine, 17. Urea, 19. Fermentation, 20. Uric Acid and 
Urates, 22. Kreatinin, 23. Hippuric Acid, 25. Origin of Hippuric Acid, 
25. Indican, 26. Origin of Indican, 27. Urobilin, 27. Oxalic Acid, 29. 
Benzoic Acid, 29. Origin of Benzoic Acid 29 



CHAPTER II. 

Inorganic Constituents of Urine, 31. Separation of Inorganic Bodies from 
Urine, 31. Chlorine, 32. Phosphoric Acid, 32. Basic, Neutral, and 
Acid Phosphates, 32. Deportment of Acid Phosphates with Alkalies, 33. 
Deportment of Soluble Phosphates with Soluble Salts of Calcium and Mag- 
nesium, ^^. Deportment of Basic and Neutral Phosphates with Acids, 34. 
Glycerin- Phosphoric Acid, 34. Sotnischewsky's Method for Testing for 
Glycerin-Phosphoric Acid in Urine, 34. Phosphorus Compounds in Healthy 
and Diseased Urine, 35. Sulphur Compounds, 36. Deportment of Sul- 
phuric Acid in Sulphates and in Ester Compounds, 36. Sulphur not in 
Sulphates and Esters, 36. Origin of Sulphur Compounds in Urine, 37. 
Carbonic Acid, 38. Compounds of Calcium and Magnesium, 38. Ammonia, 
39. Origin of Ammonia in Urine, 39. Potassium and Sodium 40 



CHAPTER III. 

Bodies in Diseased Urine, 42. Albuminous Bodies, including Mucine, 42. 
Serum-Albumen, 42. Globuline and Hemialbumose, 43. Peptone, 43. 
Mucine, 44. Color Tests for Albuminous Bodies, 44. Millon's Reagent, 
44. Table of Reactions of Albuminous Bodies, including Mucine, 45. 
Table of Color Tests, 46. Methods of Determining the Presence of Albu- 
minous Bodies, 46. The Nitric Acid Test, 46. The Acetic Acid and 
Sodium Chloride Test, 46. Determination of the Presence of Hemialbumose, 
47. Peptone, 47. Globuline and Serum-Albumen, 47. Mucine, 48. 
Diabetic Sugar, 49. Tr'ommer's Test, Modified by Salkowski, 50. Test 
with Fehling's Solution, 50. The Fermentation Test, 52. The Phenyl- 
hydrazin Test, 53. Methods for Separating Small Quantities of Sugar from 
Urine, 54. Detection of Sugar in Albuminous Urine, 55. Inosit, 55. Test 
for Inosit in Urine 55 

vii 



VI 11 CONTENTS. 



CHAPTER IV. 



Bodies in Diseased Urine Continued, 57. Biliary Acids, 57. Pettenkoffer's 
Test for Biliary Acids, 57. Separation of Biliary Acids from Urine, 58. 
Coloring Matters of the Bile in Urine, 58. Tests for Biliary Coloring 
Matters in Urine, 59. Coloring Matters of the Blood, 60. The Spectro- 
scope, 62. Spectroscopic Test for Haemoglobin and its Derivatives, 62. 
Heller's and Struve's Tests for Hsematin in Urine, 63. Blood in Urine, 63. 
Blood Corpuscles, 64. Fibrin, 64. Leucin, 64. Separation of Leucin 
from Urine, 64. Tyrosin, 65. Test for Tyrosin in Urine, 65. Fat, 66. 
Tests for Fat in Urine, 66. Dreschel's Apparatus for Extraction of Fats, 
67. Cholesterin and Lecithin, 67. Separation of Cholesterin from Urine, 
67. Tests for Neurin and Glycerin- Phosphoric Acid produced from the 
Action of Barium Hydrate on Lecithin 68 



CHAPTER V. 

Sediments in Urine 70 

Sediments Peculiar to Urine of Strong Acid Reaction, 70. The Acid Urates 
and Uric Acid, 70. Hippuric Acid, 71. Calcium Sulphate, 72. Sediments 
Peculiar to Urine of Strong Alkaline Reaction, 72. Calcium and Magne- 
sium Phosphates and Magnesium Amnion. Phosphate, 72. Neutral Calcium 
Phosphate, 73. Ammon. Acid Urate, 74. Calcium Oxalate, 74. Calcium 

Carbonate 75 

Sediments not depending on Reaction of the Urine 76 

Cystin, 76. Separation of Cystin from Urine when in Solution, 76. Cystin as 
a Sediment, 76. Tyrosin, 77. Epithelial Casts, 77. Hyaline Casts, 77. 
Waxy Casts, 78. Blood Casts, 79. Epithelial Cells, 79. Blood Corpus- 
cles, 80. Pus, 80. Spermatozoa, 82. Bacteria, 82. Sarcina and other 
Microorganisms 83 



CHAPTER VI. 

Scheme for the Qualitative Analysis of Healthy and Diseased Urine, 85. Sedi- 
ments, 88. Microscopic Examination of Sediments, 89. Staining, 90. 
Scheme for the Qualitative Analysis of Sediments 91 



CHAPTER VII. 

Concretions or Stones in the Bladder, 95. Constitution and Physical Properties 
of Concretions, 95. Some of the Causes of the Formation of Concretions, 
95. Scheme for the Qualitative Analysis of Concretions or Stones 96 



CONTENTS. IX 

QUANTITATIVE ANALYSIS. 

CHAPTER VIII. page 

Filter Paper and Filtering, 101. Evaporating, 102. Drying, 103. Ashing Fil- 
ters and Heating Precipitates, 103. Chemical Balance and Weights, 104. 
Weighing, 105. Vessels Required for Measuring Fluids, 106. Desiccators, 
Tongs, Crucibles, etc., 108. Preparation of Solutions for Volumetric Analy- 
sis, 108. Normal Oxalic Acid, 109. Normal. Potassium Hydrate, 109. 
Normal Sulphuric Acid, 112. Normal Hydrochloric Acid, 113. Solution 
of Litmus, 114. The Barium Mixture, 115. The Magnesium Mixture, 115. 
Millon's Reagent 115 



CHAPTER IX. 

The Quantity of Urine Passed in 24 hours, 116. Specific Gravity, 116. Solids 
of the Urine, 117. Inorganic Substances, 117. Coloring Matter, 118. 
Acidity of the Urine, 118. Urea, 119. Liebig's Method, Modified by 
Pfliiger, 119. Estimation of the Quantity of Urea in Diseased Urine by 
Liebig and Pfliiger's Method, 125. Knop's Method, Modified by Greene... 125 



CHAPTER X. 

Total Quantity of Nitrogen in Urine, 128. Dumas' Method, 128. Varrentrapp 
and Will's Method, 131. Kjeldahl's Method, 134. Remarks on the 
Methods of Estimating the Quantity of Urea and Nitrogen in Urine, 137. 
Uric Acid — Salkowski's Method, Modified by E. Ludwig, 138. Kreatinin — 
Neubauer's Method, Modified by Fiirbringer, 139. Oxalic Acid — Neu- 
bauer's Method, Modified by Fiirbringer 141 



CHAPTER XL 

Phosphoric Acid of the Phosphates, and of Glycerin- Phosphoric Acid, 143. Neu- 
bauer and Zuelzer's Method, 143. Phosphoric Acid of the Phosphates, 145. 
Total Quantity of Phosphoric Acid, 146. Sulphur Compounds, 147. Sul- 
phuric Acid or Sulphur in Sulphates and Ester Compounds — The Gravime- 
tric Method, 147. The Volumetric Method (Wildenstein, Briiggelmann, and 
Neubauer), 149. Sulphuric Acid in the Ester Compounds, 152. Sulphur 
not in Sulphates or Ester Compounds, 152. Method of Estimating — total 
Quantity of Sulphur in Urine, 152. Chlorine — Volhard's Method, Modified 
by Salkowski, 153. Neubauer's Method, 156. Remarks on the Methods 
of Estimating the Quantity of Chlorine in Urine, 157. Potassium and 
Sodium, 157. Ammonia — Schlosing and Neubauer's Method, 159. Cal- 
cium and Magnesium Oxides, 161. Calcium Oxide— The Gravimetric 
Method, 161. The Volumetric Method (Neubauer), 162. Magnesium 
Oxide — The Gravimetric Method, 163. The Volumetric Method (Neu- 
bauer) 164 



CONTENTS. 



CHAPTER XII. 



Albumen — Scherer's Method, 166. Globuline — Hammarsten's Method, 167. 
Pohl's Method, 168. Hemialbumose — The Gravimetric and Optical 
Methods, 169. Peptone — Hofmeister's Method, 169. Remarks, 170. 
Sugar — Fehling's Method, 170. Fehling's Method, Modified by Pavy, 173. 
Roberts' Method, 175. The Optical Method, 175. Remarks on the 
Methods of Estimating the Quantity of Sugar in Urine 180 



APPENDIX. 

TABLE I. 
Nitrogenous Compounds, except Urobilin, in Normal Urine 181 

TABLE II. 

Number of Parts of Nitrogen and other Constituents of Elements of Food in 

1000 Parts — From Zuelzer 182 

TABLE III. 

Relative Number of Parts of Constituents of Certain Animal and Vegetable 

Bodies for 100 Parts of Nitrogen which they contain— From Zuelzer 182 

TABLE IV. 

Indicating Changes in the Constitution of the Urine by the Ingestion of 

Different Kinds of Food — From Zuelzer 183 



ERRATUM. 
On page 63, 12 lines from top, " crystals of hjematin " should read " crystals of heemin. 



LIST OF ILLUSTRATIONS. 



PAGE 

Table I . Spectra of Coloring Matters 

of the Blood, etc 60 

Figure 1. Kreatinin Zinc Chloride... 24 

' : 2. Fermentation Tube 52 

3. Spectroscope 61 

4. Hsemin (Haematin hydro- 

chlorate) 63 

" 5. Leucin and Ty rosin 65 

" 6. Fat 66 

" 7. Dreschel's Apparatus for 

Extraction of Fats 67 

" 8. Urates and Uric Acid 71 

" 9. Uric Acid Separated by 

an Acid 71 

" 10. Hippuric Acid 71 

" II. Calcium Sulphate, from 

Furbringer 72 

" 12. Calcium and Mag. Phos- 
phates (Basic) 72 

" 13. Mag. Ammon. Phosphate.. 72 
' 14. Calcium Phosphate (Neu- 
tral). From Peyer 74 

'• 15. Ammon. Acid Urate 74 

li 16. Calcium Oxalate 74 

'• 17. Calcium Carbonate 76 

" 18. Cystin 76 

" 19. Epithelial Casts. From 

Salkowski 78 

" 20. Hyaline Casts. From 

Peyer 78 



Figure 21. 
" 22. 

" 23. 

" 24. 



25- 

26. 
27. 

28. 



33- 
34- 
35- 

36. 

37- 

38. 



PAGE 

Waxy Casts. From Peyer 78 
Blood Casts. From 

Peyer 79 

Epithelial Cells, from Uri- 

niferous Tubes 79 

Epithelial Cells, from 

Bladder 79 

Epithelial Cells, from 

Neck of Bladder 79 

Blood Corpuscles. 81 

Pus Corpuscles 81 

Spermatozoa 81 

Micrococcus Ureae with 
crystals of Mag. Ammon. 
Phos. and Ammon 

Urate 83 

Sarcina 83 

Penicilium Glaucum 

(Mould) 83 

Apparatus for Estimation 

of Urea ... 126 

Zulkowsky's Azotometer. 130 

Syphon Filter 150 

Apparatus for Estimation 

of Ammonia 160 

Wild's Saccharimeter 176 

Views in Wild's Sacchari- 
meter 177 

Views in Laurent's Saccha- 
rimeter 179 



CHEMICAL ANALYSIS 



OF 



THE URINE 



CHAPTER I. 



Properties of the Urine — Nitrogenous Bodies, including Oxalic and Benzoic Acids — 
Urea — Fermentation — The Xanthin Group of Compounds — Uric Acid — Kreatinin 
— Hippuric Acid — Indican — Urobilin — Oxalic Acid — Benzoic Acid. 

PROPERTIES OF THE URINE. 
Normal urine, when fresh, is generally clear, and after the lapse 
of a few hours a cloudy or flaky sediment in irregular masses is 
usually observed at the bottom of the vessel containing the urine. 
As urine generally holds in solution acid phosphates and sul- 
phates it is usually acid in reaction, but it is sometimes neutral 
or alkaline, depending very much on the diet. If the urine is 
alkaline when fresh it is usually cloudy or opaque, as there are 
certain compounds of calcium and magnesium which are insol- 
uble in alkaline fluids. The reason that different kinds of food 
affect the reaction of the urine is that in meat, eggs, cheese, etc., 
there are considerable quantities of phosphorus and sulphur and 
less of the metals of the alkalies — potassium and sodium. In 
the urine phosphorus and sulphur combined with oxygen and 
metals form acid salts if the metals are not in sufficient quantity 
to form basic salts. On - the other hand, in vegetable food the 
quantity of potassium is greater, and by the ingestion of food 
largely vegetable in kind the blood becomes more alkaline in 
reaction than by the ingestion of animal food. 
2 17 



18 CHEMICAL ANALYSIS OF THE URINE. 

When normal urine is alkaline the urine formed subsequently 
is rendered acid by the ingestion of a dilute acid. No free acid, 
however, is formed in the urine, but the basic or neutral phos- 
phates are changed to acid salts — 

Ca 3 (P0 4 ) a -f 4 HC1 == 2CaCl 2 + CaH 4 (POJ 2 , 

Basic Phosphate. Acid Phosphate. 

or 2CaHP0 4 + 2 HC1 = CaCl 2 + CaH 4 (P0 4 ) 2 . 

Neutral Phosphate. Acid Phosphate. 

If dilute acid be administered in excess to a carnivorous animal, 
an increased quantity of ammonia combined with acids is found in 
the urine. The ammonia is formed at the expense of the quan- 
tity of urea. After a hearty meal the urine is usually alkaline 
until the lapse of a few hours, when the urine formed is acid, 
unless the food is largely vegetable in kind. During the period of 
secretion of the gastric juice the blood becomes less alkaline as it 
loses chlorine, which leaves bases to form hydrochloric acid in the 
gastric juice ; consequently, the urine becomes neutral or alkaline 
from the presence of the alkaline carbonates. Later in the period 
of digestion, when absorption and assimilation take place, this 
drain from the blood ceases, as the hydrochloric acid is, in part, 
absorbed, and part combines with the potassium and sodium of 
the biliary salts, and some of the phosphorus and sulphur of the 
food appear in the urine in acid combinations. 

Normal urine varies greatly in color, from being nearly color- 
less to dark brown. Usually, however, it is of an amber or straw 
color. In disease, the urine is generally more highly colored 
than in health. Exceptions are in diabetes mellitus, chlorosis, 
anaemia, etc. In the febrile state the urine is highly colored, with 
high specific gravity, owing to increased quantity of urea. Color- 
ing matters of the bile or blood may impart a deep color — yellow, 
red or dark brown — without there being an increased quantity of 
urea ; or the urine may be of an abnormal color, due to the inges- 
tion of rhubarb, tarry substances, etc., and the urine be normal 
in every other respect. As the color of normal urine varies 
greatly, so its odor is subject to variation. The chemical con- 
stitution of the body to which the urine owes its odor is not 
known. It volatilizes very slowly, as urine does not lose all of 
its odor by boiling or evaporating. 

The specific gravity of the urine varies from 1002 to 1030; 



UREA. 19 

that is, a volume of urine would weigh ioio or 1025 grammes 
or grains, while an equal volume of water would weigh 1000 
grammes or grains. The specific gravity of urine is lessened by 
drinking much water, while it is increased by a warm, dry atmos- 
phere and consequent free perspiration. As activity of the skin 
is increased by exercise to exhaustion, so the specific gravity of 
the urine is likewise increased from the formation of an increased 
quantity of urea. On the other hand, by moderate exercise the 
specific gravity of the urine is not increased. 

UREA. 

Of the constituents of urine containing nitrogen, urea is the 
most important, as it constitutes nearly 3 per cent, of the weight 
of urine. 

It is composed of carbon, hydrogen, oxygen and nitrogen and 

has the formula of 

CH 4 N 2 0, 

which group of elements has the structural formula of 



CO 



/NH 



\NH 2 ' 

It is a substitution compound of ammonia, NH 3 , in which one 
atom of hydrogen is substituted by CO, as 



NH 2 \ 

NH 2 / LUl 



It is apparent that urea is an organic body, yet the structure of 
the molecule is comparatively simple, being very little more com- 
plex than that of an inorganic molecule. Urea is the principal 
decomposition product of muscular and other tissues. It is 
formed in the blood (Schmiedeberg) by the withdrawal of the 
elements of water from ammonium carbonate — 
(NH 4 ) 2 C0 3 — 2H 2 = CH 4 N 2 0. 

Ammon. Carb. Urea. 

46.66 per cent, of urea is nitrogen. The average quantity of 
urea excreted by the kidneys of an adult in twenty-four hours 
is 31 grammes. 

One gramme of urea corresponds to 13.72 grammes muscu- 
lar tissue. By consulting Table 1 in the Appendix it is seen that 
in 31 grammes urea there are 14.464 grammes nitrogen, and 
about 93 per cent, of the total quantity of nitrogen excreted 
by the kidneys is in this compound ; but the quantity of urea 



20 CHEMICAL ANALYSIS OF THE URINE. 

excreted in twenty-four hours is subject to variation, ranging from 
25 to 40 grammes, depending not only on the quantity of the nitro- 
genous elements of food having entered the circulation, but on 
the rate of the desassimilation of tissues, and as the latter process 
is facilitated by fever, urea appears in increased quantity in the 
febrile state. In determining the rate of desassimilation of the 
tissues by the quantity of urea excreted, the kind and quantity of 
food ingested and the quantity of nitrogen in the faeces are taken 
into account; hence simple estimations of urea in the urine do not 
afford sufficient data, but in fevers there is wasting of the tissues, 
with a corresponding increase in quantity of urea excreted and 
diminished quantity of sodium chloride or chlorine in the urine. 
On the other hand, when there is no abnormal waste of tissues, but 
there is an increased quantity of urea excreted coming from the 
nitrogenous elements of food ingested, the quantity of chlorine (in 
combination) in the urine is likewise increased. In diabetes and 
anaemia the quantity of urea excreted is increased without fever. 
Urea is a colorless crystalline body ; the crystals contain no water 
of crystallization and undergo no change in the air. It is soluble 
in water and alcohol and the solution is neutral in reaction. At 
130 urea melts, if free of water. It acts as a base, forming salts 
with acids. Urea can be separated from the urine by adding 
barium mixture (for the preparation of which refer to Chapter 
vin) to the urine — one or two litres — until, after mixing well by 
stirring with a glass rod, a precipitate ceases to form. Filter, and 
evaporate the filtrate on a water bath to syrupy consistence. By 
standing twenty-four hours the urea will crystallize, when the mass 
is pressed between porous paper, transferred to a litre flask and 
treated with about 500 c.c. 90 per cent, alcohol, and after shaking 
more or less for thirty minutes, filter, and evaporate the filtrate 
on a water bath. The residue is nearly pure urea, but is highly 
colored by the coloring matter of the urine, to remove which dis- 
solve in water and treat with a solution of potassium perman- 
ganate ; filter, and evaporate the filtrate on a water bath to 
dryness. 

FERMENTATION. 
Normal urine, when exposed to the air at ordinary tempera- 
tures, undergoes remarkable changes: losing its acidity, it becomes 
turbid, emits the uriniferous odor, and when heated gives off 



THE XANTHIN GROUP OF COMPOUNDS. 21 

alkaline vapors. By microscopic examination the urine is found 
impregnated with bacteria, the Micrococcus urece. Besides bacteria, 
if the urine is strongly alkaline in reaction, crystals of magne- 
sium, ammonium phosphate, ammonium urate, a and b, Fig. 29, 
page 83, and calcium oxalate, Fig. 16, page 74, are found in the 
sediment. There is in solution a large amount of ammonium 
carbonate, urea having been decomposed by absorbing water — 
CH 4 N 2 + 2H 2 = (NH 4 ) 2 C0 3 . 

Urea. Ammon. Carb. 

That bacteria is the cause of the fermentation is shown by 
putting a small quantity of fermenting urine into fresh urine, 
when the latter will ferment much sooner than it otherwise 
would. It is, therefore, of great practical importance to put 
fresh urine, for analytical purposes, into clean vessels, and for 
closing bottles or flasks containing urine to employ corks which 
have not been in use for the same purpose. Obstinate diseases 
of the bladder are sometimes produced by introducing the 
micrococcus ureae into the bladder by the use of catheters which 
had been used before, but not properly cleaned. In case of fer- 
mentation taking place in the bladder, the alkaline reaction of 
the urine is generally due to the formation of ammonium car- 
bonate, but occasionally bacteria is found in fresh urine decidedly 
acid in reaction, the process not having continued long enough 
for the formation of sufficient ammonium carbonate to transform 
the acid salts to basic compounds. 

Urine undergoing fermentation is turbid in every part, while 
urine alkaline from the presence of potassium and sodium car- 
bonates, by transmitted light, appears more or less clear, with 
floating particles or suspended flakes. 

To prevent urine from fermenting a few days, it is kept in 
bottles surrounded by ice, or, in the absence of which, it is ren- 
dered strongly acid with hydrochloric acid. 

THE XANTHIN GROUP OF COMPOUNDS. 

Sarkin or Hypoxanthin, C 5 H 4 N 4 0. 
Xanthin, C 5 H 4 N 4 2 . 

Uric Acid, C 5 H 4 N 4 3 . 

The bodies here enumerated, with another member of the 

group, guanin — 

a.H e N e O, 



22 CHEMICAL ANALYSIS OF THE URINE. 

are found in glandular and muscular tissues. It is seen by the 
formulae of these bodies that they are closely related. That 
in all probability they have a common origin — products of the 
reduction of albuminous bodies — is rendered more evident by 
the fact that each has been produced from other members of the 
group by chemical processes. Xanthin is found in the urine in 
exceedingly small quantities. Refer to Table I in the Appendix. 
Whether sarkin is a constituent of normal urine is doubtful. A 
body resembling it in many respects is found in the urine, but in 
mere traces. 

URIC ACID. 
Although uric acid is found in human urine in small quan- 
tity, yet it is a constant constituent. In the urine of birds and 
reptiles it is in great quantity. 33.3 per cent, of uric acid is nitro- 
gen. It is more complex in constitution than urea. Its for- 
mula is — 

C 5 H 4 N 4 3 . 

Usually uric acid is in combination in the urine, forming 
urates, yet it sometimes appears as a sediment. In twenty-four 
hours 0.2 to 1 grm. uric acid is excreted by the kidneys. The 
average quantity is about 0.6 grm. Consult Table I in the Appen- 
dix. When the food is composed principally of meats, eggs, 
etc., the quantity of uric acid excreted is increased. As the tissues 
are reduced or desassimilated in far greater quantity during fever 
than during health, the amount of uric acid excreted is likewise 
increased. In health the relative quantity of uric acid and 
urea is 1 to 51. In fever attended by checked aeration of 
the blood, as in pneumonia, the relative quantity of uric acid 
is increased. An increase of uric acid is likewise observed 
in leucocythsemia. In this disease aeration of the blood is 
diminished by reason of the diminution of the red corpuscles 
of the blood. In articular rheumatism the quantity of uric 
acid excreted is not increased ; the sediment of the urates 
and uric acid, so often observed in this disease, results from 
concentration of the urine and strong acid reaction. In dia- 
betes and advanced stages of Bright's disease the quantity of 
uric acid in the urine is diminished. 

Uric acid is a crystalline body, rhombic and tabular in form. 
It is tasteless, odorless, and in cold water nearly insoluble. Uric 



KREATININ. 23 

acid forms salts with potassium, sodium, or ammonium carbonate 

or hydrate, according to the equations — 

C 5 H 4 N 4 3 -f- 2NaOH = C 5 H 2 Na 2 N 4 3 -f 2H 2 0, 
C 5 H 4 N 4 3 + Na 2 C0 3 .= C 5 H 3 NaN 4 3 + NaHC0 3 . 

From the formulae of the urates formed by these reactions, it 
is seen that there are two different urates, one containing one 
atom of Na or K, and the other two atoms ; consequently, 
uric acid is a dibasic acid, that is, it contains two atoms H, 
which may be substituted by two atoms of a monad metal. 
Urates containing but one atom of K, Na or NH 4 are acid urates, 
those containing two are neutral urates. There are also urates 
composed of a molecule acid urate, and a molecule uric acid, as — 
C 5 H 3 KN 4 3 , C 5 H 4 N 4 3 - 

The acid urates are frequently found as a sediment in urine of 
acid reaction. All urates are decomposed by dilute hydrochloric 
acid, with the liberation of uric acid — 

C 5 H 2 Na 2 N 4 3 + 2HC1 = 2NaCl + C 5 H 4 N 4 3 . 

To separate uric acid from the urine, it is concentrated by 
evaporation to about one-half its volume, when it is rendered 
strongly acid with dilute hydrochloric acid, and, after standing 
twenty-four hours, crystals of uric acid are observed adhering to 
the walls of the glass containing the urine — sedimentum lateri- 
tium. By filtering and collecting the crystals on the filter and 
washing with some cold water, uric acid can be obtained nearly 
pure but highly colored. As the acid urate of ammonium is far 
less soluble in water than the corresponding urates of sodium or 
potassium, it forms when a solution of ammonium chloride is 
added to the urine, especially after the lapse of a few hours. 

Besides the microscopic examination showing crystalline forms 
of uric- acid and urates, Fig. 8, page 71, its presence can readily 
be determined by evaporating some strong nitric acid, mixed with 
uric acid on a watch glass, to dryness, and treating the residue 
with a small quantity of ammonia water, when a purple-red color 
is observed. 

- KREATININ. 

Kreatinin, C 4 H 7 N 3 Q, and kreatin, C 4 H 9 N 3 2 , differ, as seen by 
their formulae, the latter containing H 2 more. Both are con- 
stituents of urine in certain conditions, as these' bodies change 



24 CHEMICAL ANALYSIS OF THE URINE. 

into each other, according to the condition. In alkaline urine, 
kreatin is in greater quantity. In urine strongly acid, kreatinin 
is found with either no kreatin or a mere trace. Physiologically, 
they may be regarded as one body, but, as the urine is usually 
acid, kreatinin is considered the normal constituent. 

Chemically, it is regarded as a substitution compound of ammo- 
nium, as it is a base, and, like ammonia, it forms salts by direct 
union with acids without the separation of water. In the urine 
of an adult secreted in twenty-four hours, kreatinin is in quan- 
tities from a trace to 1.3 grm.; the average is about 0.7 grm. In 
fevers it is increased in quantity, and in diabetes the quantity is 
diminished. To determine the presence of kreatinin in the urine 
(Neubauer and Salkowsky), 500 c.c. urine is treated with milk 
of lime until it becomes alkaline in reaction, when a solution of 
calcium chloride is added until a precipitate ceases to form, known 
by filtering a small quantity and testing the filtrate with a solution 
of calcium chloride, and if no precipitate forms, enough has been 
added. Calcium hydrate and chloride serve the purpose of 
separating the phosphoric acid. Filter, and render the filtrate 
distinctly acid with hydrochloric acid, avoiding an excess of acid. 
Evaporate nearly to dryness, and, when cold, treat with about 
100 c.c. strong alcohol. Transfer the residue and fluid to a flask 
or beaker, render alkaline with a solution of sodium carbonate; 
after which render acid with acetic acid. The solution should 
be acid in reaction, but not with hydrochloric acid.. After stand- 
ing ten hours, filter, and to the filtrate add a strong alcohol solu- 
tion of zinc chloride, to which some sodium acetate is added, if 
the solution is acid in reaction. Kreatinin zinc chloride will 

form. The formula is — 

(C 4 H 7 N 3 0) 2 ZnCl 2 . 

Fig. i. The microscopic character of this body is shown by 

Fig. 1. If the urine contains sugar, it is decom- 
posed by putting some yeast in it and keeping it at 
25 ° to 30 twenty-four hours. Albuminous urine is 
rendered acid with dilute sulphuric acid, boiled 
twenty minutes and filtered before it is subjected to 
further treatment. 
Kreatinin, when in aqueous solution, imparts the alkaline reac- 
tion. With acids it combines, forming salts. When a solution 




HIPPURIC ACID. 25 

of sodium nitroprussiate is added to a dilute solution of kreatinin 
(Weyl's test), and then sodium hydrate is added, the solution 
changes to a red color, which, by standing, turns yellow. This 
is the most delicate test for kreatinin in urine. The presence of 
sugar or albumen in the urine does not interfere with the reaction. 
Kreatinin reduces CuO in Fehling's solution to Cu 2 0. 

HIPPURIC ACID. 

In the urine of herbivorous animals hippuric acid is found in 
considerable quantities. In human urine the quantity is less. In 
the urine formed in twenty-four hours by a man of average weight 
there is 0.15 to 0.6 grm. hippuric acid. The average quantity is 
0.4 grm. The quantity of hippuric acid excreted by the kidneys 
depends very much on the kind of food, yet with a nitrogenous 
(animal) diet the acid does not disappear from the urine. It has 
been found that when albumen is decomposed by means of the 
pancreatic ferment, phenylpropionic acid is formed, and, by oxi- 
dation, this acid forms benzoic acid, which, by uniting with glyco- 
col, produces hippuric acid. The following equations will serve 
to illustrate some of the chemical changes which take place in 
the formation of benzoic and hippuric acids : — 

C 6 H 3 — CH 2 — CH 2 — COOH + 60 = 2H,0 + 2CO, + C 6 H 5 — COOH. 

Phenylpropionic Acid. Benzoic Acid. 

C 6 H 5 _ COOH + CH ,<^ H = CH 2 /£H- C 6 H 5 - CO + ^ 

Benzoic Acid. Glycocol. Hippuric Acid. 

Glycocol, as well as phenylpropionic acid, is formed in the 
process of intestinal digestion. Phenylpropionic acid having 
entered the circulation is oxidized, forming benzoic acid as one 
of its products, when the latter acid, uniting with glycocol, forms 
hippuric acid with the separation of water. This is the source of 
hippuric acid in the urine when nitrogenous food is taken, but, as 
the quantity of the acid is greatly increased by a diet composed 
principally of vegetables, it cannot be considered the only source. 
As hippuric acid originates in different ways, it is necessary to 
take into account many data in determining if it arises from 
pathological processes. ' 7.87 per cent, hippuric acid is nitrogen. 
It is a crystalline body, usually crystallizing in four-sided prisms. 
The acid is colorless and has a bitter taste. It is not very soluble 
in cold water, but more soluble in hot water. In alcohol it is 



26 CHEMICAL ANALYSIS OF THE URINE. 

soluble. Hippuric acid yields but few characteristic reactions, so 
that its separation from the urine is necessary before subjecting 
it to reactions. For this purpose about iooo c.c. urine (Bunge 
and Schmiedeberg) are rendered alkaline with sodium carbonate, 
filtered, and the filtrate evaporated to near dryness, and the residue 
treated with alcohol in several portions of 50 c.c. and filtered each 
time. Evaporate the alcohol and treat the residue with a small 
quantity of water, transfer to a glass cylinder, acidify the mixture 
with hydrochloric acid and shake up five times with about 30 c.c. 
acetic ether, and each time, when the ether separates from the 
water, it is removed by means of a syphon or pipette. The acetic 
ether is washed by shaking it up with water in a glass cylinder, 
and drawing off the acetic ether as before. By evaporating the 
acetic ether to dryness at ordinary temperatures, crystals of hip- 
puric acid will be found in the residue. To remove benzoic acid 
and any fat that may be present, dissolve the residue in water and 
transfer to a glass cylinder, shake up with benzole, and when it 
has separated from the water, remove by means of a syphon or 
pipette. By evaporating the aqueous solution, hippuric acid will 
separate in the form of crystals. Besides the crystalline forms of 
hippuric acid, Fig. 10, page 71, its presence is determined by 
treating it with strong nitric acid and evaporating on a water bath 
to dryness, when the residue is transferred to a dry test tube and 
carefully heated. If hippuric acid is present, a peculiar odor, 
resembling that of the oil of bitter almonds, is produced, due to 
the formation of nitrobenzole — C 6 H 5 N0 2 . 

INDICAN. 
Indican is the potassium salt of a substitution compound of 
sulphuric acid, H 2 S0 4 , in which the group of elements C 8 H 6 N 
takes the place of one atom of hydrogen of H 2 S0 4 . The free 
acid, indoxylsulphuric acid, 

C 8 H 6 N\ S0 

is not stable, but its salts are well known. Indican is not a 
coloring matter, but when indoxylsulphuric acid is oxidized in- 
digo blue is formed. 

2C 8 H 6 N\ S0 ^ + q 2 = 2 h 2 S0 4 + C 16 H 10 N 2 O 2 . 

Indoxylsulphuric Acid. Indigo Blue. 



UROBILIN. 27 

The formula of indol is C 8 H 7 N. It is formed in the intestines by 
the action of pancreatic juice and bacteria on albuminous bodies. 
Much of it passes off with the faeces, and that which is absorbed 
appears in the urine as indican. It is known that this is one 
source, if not the only one, of indican in the urine, for when indol 
is introduced into the blood, the quantity of indican in the urine 
is greatly increased. When the conditions for increased ferment- 
ation of the contents of the intestines are favorable, as in obstinate 
constipation and peritonitis, the quantity of indican in the urine 
is greatly increased, and when the quantity of albuminous food 
is reduced the amount of indican in the urine is likewise reduced. 
About 1 1 milligrammes indican is excreted by kidneys of a man 
of average weight in twenty-four hours. Indican is a solid, soluble 
in water and partly soluble in alcohol. Indigo blue when brought 
in contact with nascent hydrogen changes to indigo white — 
C 16 H 10 N 2 O 2 + H 2 = C 16 H 12 N 2 2 . 

Indigo Blue. Indigo White. 

Indigo blue is insoluble in water ; soluble in alcohol, ether and 
chloroform. The test for indican in the urine is based on the 
fact that indigo blue is formed by the oxidation of indoxylsul- 
phuric acid. 

To one part urine (Jaffe's test) add one part hydrochloric acid, 
specific gravity 1.124. The acid serves the purpose of decom- 
posing indican, liberating indoxylsulphuric acid — 

CsH6 K/ S0 4 + HC1 = KCl + C8H6 h/ S0 4- 

Indican. Indoxylsulphuric Acid. 

To the fluid add, drop by drop, a filtered solution of bleaching 
powder (1 part to 20 parts of water). Shake the mixture well 
after each addition of the solution, and when a decided change of 
color of the fluid takes place shake up with a few cubic centi- 
metres of chloroform. The latter as it settles will be colored blue. 

Besides indican there are in the urine, under certain circum- 
stances, analogous compounds in which phenyl, C 6 H 5 , and cresyl, 
C 7 H 7 , are substituted for one atom of hydrogen of H 2 S0 4 . Refer 
to sulphuric acid, page- 36. 

UROBILIN. 

The normal color of the urine is not due to any one coloring 
matter as found by spectroscopic examination. The most im- 



28 CHEMICAL ANALYSIS OF THE URINE. 

portant coloring matter, however, is urobilin. The formula of 
urobilin is C32H 40 N 4 O 7 . It is a derivative of bilirubin, a consti- 
tuent of bile. This body, by the action of nascent hydrogen, 
changes to urobilin — 

2C 16 H 18 N 2 3 +' H 2 + H 2 = C 32 H 40 N 4 O 7 . 

Bilirubin. Urobilin. 

In the intestines nascent hydrogen is formed, and by it bili- 
rubin of the bile is reduced to urobilin, and the latter is in 
part absorbed by the blood and excreted by the kidneys. There 
is reason to believe that urobilin is itself in part reduced by 
nascent hydrogen, from the fact that urine sometimes becomes 
more deeply colored by exposure to the air, by which a colorless 
body is oxidized to form urobilin. This has been more satis- 
factorily proven by separating the coloring matters from the 
urine by shaking with some basic lead acetate, filtering, and 
exposing the colorless filtrate to the air, when it may become 
colored. By spectroscopic examination of the filtrate, when col- 
ored by oxidation, urobilin is found. Urobilin is a dark, amor- 
phous powder. It is nearly insoluble in water, soluble in alcohol, 
ether, and chloroform. In a solution of potassium, sodium, or 
ammonium hydrate it dissolves. 

From alkaline solutions, rendered nearly neutral, it is precipi- 
tated by a salt of a heavy metal, for example, lead acetate, silver 
nitrate or copper sulphate. It is partly precipitated from alkaline 
solutions by rendering them acid. Solutions of urobilin, when 
examined by the spectroscope, exhibit a band between the lines 
b and F, Spectrum 4, Table i,page 60. For spectroscopic exami- 
nation, prepare a solution in ammonium hydrate to which some 
zinc chloride has been added. The band is brought out more 
distinctly by these solvents. To test for urobilin, the use of the 
spectroscope is more satisfactory than the employment of chem- 
ical tests. For description and use of the spectroscope refer to 
page 62. As there is a very small quantity of urobilin in normal 
urine, the urine of a person having fever is examined for urobilin. 
About 200 c.c. of the urine is treated with basic lead acetate, fil- 
tered, and the precipitate is washed once with cold water and dried 
at a low temperature. Triturate the dry mass in a mortar with 
strong alcohol (20-25 c.c), to which add some dilute sulphuric 
acid to separate the lead, and after standing a few hours filter, 



BENZOIC ACID. 29 

render the filtrate alkaline with ammonium hydrate, add a small 
quantity of a solution of zinc chloride and examine the solution 
with the spectroscope. 

OXALIC ACID. 
Oxalic acid combined with potassium and calcium is found in 
many vegetables. The formula of it is C 2 H 2 4 . It is a dibasic 
acid soluble in water. The oxalates of potassium, sodium, and 
ammonium are soluble in water. Calcium oxalate is insoluble 
in water and dilute acetic acid, but soluble in water containing 
hydrochloric acid. Calcium oxalate crystallizes with one mole- 
cule water in tabular form and with three molecules water in 
octahedral forms; the latter usually form slowly, while the former 
results by the crystallization taking place in a short time. As 
normal urine is usually acid, calcium oxalate is held in solution. 
It is only when the urine becomes neutral or alkaline that it 
appears as a sediment. The changing of the urine from the acid 
to the neutral or alkaline condition is usually a slow process, so 
that calcium oxalate usually appears as a sediment in the form 
of octahedral crystals. For the character of the crystalline form 
refer to Fig. 16, page 74. To determine the presence of oxalic 
acid in the urine, about one litre urine is treated with an excess 
of a solution of calcium chloride and rendered alkaline with 
ammon. hydrate. After the lapse of a few hours the mixture is 
rendered acid with acetic acid, avoiding more than necessary. 
The acetic acid dissolves the phosphates of calcium and magne- 
sium, but calcium oxalate remains undissolved. The urine is 
then filtered and the precipitate is washed with water, when it is 
dissolved on the filter with a small quantity of dilute hydro- 
chloric acid, and after washing from the filter with some water 
the acid fluid is rendered alkaline with ammon. hydrate. The 
calcium oxalate will reprecipitate. After standing several hours, 
the precipitate may be examined by the microscope. 

BENZOIC ACID. 

As many kinds of vegetables, particularly fruits, contain small 

quantities of benzoic^acid, it is introduced into the blood by 

employing a vegetable diet. But whether it is introduced with 

the food, or formed from albuminous bodies in the process of 



30 CHEMICAL ANALYSIS OF THE URINE. 

intestinal digestion, it is to a great extent transformed into hip- 
puric acid by uniting with glycocol. The conditions for the 
reactions to take place are present in the kidneys, as shown by 
the experiments of Bunge and Schmiedeberg. Refer to the 
reactions above, under hippuric acid. As some of the benzoic 
acid is liable to escape this reaction, it is sometimes found in 
normal urine. By the employment of the method of Bunge and 
Schmiedeberg for the separation of hippuric acid from the urine, 
benzoic acid is also separated. The method is given above. 
Benzoic acid is separated from hippuric acid with pure benzole. 
For this purpose shake the aqueous solution in a glass cylinder 
with benzole, and, when it has separated from the water, draw it 
off with a syphon or pipette. Repeat the process two or three 
times and concentrate the benzole solution by evaporation. If 
the acid is colored and otherwise impure, it is easily purified by 
dissolving the residue of the benzole with a small quantity of hot 
water, filtering, and crystallizing out by slow evaporation. 



CHAPTER II. 

Inorganic Constituents of Urine — Chlorine, Phosphoric Acid, Basic, Neutral and 
Acid Phosphates — Deportment of Acid Phosphates with Alkalies — Deportment of 
Soluble Phosphates with Soluble Salts of Calcium and Magnesium — Deportment 
of Neutral and Basic Phosphates with Acids — Glycerin-phosphoric Acid— Phos- 
phorous Compounds in Healthy and Diseased Urine — Sulphur Compounds — 
Deportment of Sulphuric Acid as Sulphates and as Ester Compounds — Sulphur 
not as Sulphates and Esters — Origin of Sulphur Compounds in Urine — Carbonic 
Acid — Compounds of Calcium and Magnesium — Ammonia — Origin of Ammonia 
in Urine — Potassium and Sodium. 



INORGANIC CONSTITUENTS OF URINE. 
Inorganic bases in the urine are calcium, magnesium, potas- 
sium, and sodium oxides. Ammonia is also present, and is clas- 
sified as an inorganic base. There is also a small quantity of 
iron oxide in normal urine, but the quantity is so small that it is 
not considered. Inorganic acids in the urine are hydrochloric, 
phosphoric, sulphuric, and carbonic acids. Silicic acid is found 
in traces in normal urine. Inorganic bodies are separated from 
the urine by evaporating and oxidizing the residue. For this 
purpose evaporate ioo c.c. urine to dryness in a porcelain dish, 
heat gradually over the free flame until a charred mass is formed 
and vapors are no longer given off. After cooling, treat with 
small quantities of water, decant into a small filter and preserve 
the filtrate. Dry the contents of the dish, heat to a high tem- 
perature until the carbon is oxidized, when cold treat with water, 
filter, and evaporate the filtrate in the evaporating dish to dryness. 
The carbon of the residue of urine, being intimately mixed with 
the soluble bodies of the ash, is difficult to oxidize, hence they 
are removed with water before the oxidation is completed. If 
some normal urine be evaporated slowly, the crystals first formed 
are nearly pure sodium chloride, NaCl, and by continuing the 
evaporation a compound of urea and sodium chloride will crys- 
tallize out. It is well known that there is much of the sodium 
and chlorine combined in the urine, there being besides sodium 
an insufficient quantity of metals, or bases, to combine with more 
than 30 per cent, of the chlorine. Compounds, which under 
ordinary conditions are insoluble in water, are in solution in the 

31 



32 CHEMICAL ANALYSIS OF THE URINE. 

urine, owing to the formation of acid salts; thus lime and mag- 
nesia, combined with phosphoric acid, are in solution in urine of 
acid reaction. 

CHLORINE. 

The quantity of chlorine in the urine is subject to variation, as 
the kidneys are not the only channel through which the chlorides 
are eliminated from the blood. The chlorides are found in per- 
spiration, saliva, matters expectorated, bile, and the faeces. In 
health the quantity of chlorine in the urine depends largely on 
the amount of common salt taken with food, and also on the 
activity of the skin. Taking into account the amount of the 
chlorides taken with the food, the quantity of chlorine excreted 
by the kidneys during the forenoon is greater than in the same 
quantity of urine formed during the night. (Hegar.) By mental 
work the quantity of chlorine excreted is increased. During fever 
chlorine is found in the urine in less quantity than in health, but 
in convalesence from fever the quantity is generally increased, 
even if the diet remain the same. In pneumonia the quantity of 
chlorine in the urine is diminished, in diarrhoea the quantity is 
also diminished. In health the average quantity of chlorine 
excreted by the kidneys in twenty-four hours is 6 to 8 grms. 

When a soluble chloride is brought in contact with a solution 
of silver nitrate, a white precipitate is formed, AgCl. It is 
insoluble in dilute nitric acid, but soluble in ammonium hydrate. 
To test for chlorine in the urine, acidify with dilute nitric acid, 
add solution of silver nitrate; a white precipitate will form. 

PHOSPHORIC ACID IN COMBINATION AS PHOSPHATES. 

In the urine there are different phosphates of the same metal, 
according as one, two, or all three of the atoms of H of H 3 P0 4 
(phosphoric acid) is substituted by the metal. The three com- 
pounds of sodium are NaH 2 P0 4 , Na 2 HP0 4 and Na 3 P0 4 ; the first 
is known as the acid, the second the neutral, and the third the 
basic, phosphate of sodium. The formulae of the corresponding 
phosphates of calcium are — 

CaH 4 (P0 4 ) 2 , CaHP0 4 and Ca 3 (POJ 2 . 
Acid Ph. Neutral Ph. Basic Ph. 

As a calcium or magnesium atom has twice the saturating 
power of a potassium or sodium atom, the formulae differ. In 



PHOSPHORIC ACID, PHOSPHATES. 33 

the urine phosphoric acid is partly combined with sodium and 
potassium and partly with calcium and magnesium. The former 
are generally known as the phosphates of the metals of the 
alkalies, or alkaline phosphates, and the latter as the phosphates 
of the metals of the alkaline earths. The phosphates of potas- 
sium and sodium, whether acid, neutral or basic, are soluble in 
water. The acid phosphates of these metals, in solution, change 
to neutral or basic phosphates when brought in contact with potas- 
sium or sodium hydrate — 

NaH 2 P0 4 + KOH = KNaHP0 4 + H 2 0. 
NaH 2 -PC\ + 2NaOH = Na^P0 4 + 2H 2 0. 

When neutral or basic alkaline phosphates are formed in the 
urine by the action of potassium or sodium hydrate, they react 
on the calcium and magnesium in the form of acid or neutral 
phosphates or other soluble salts, precipitating them as basic or 
neutral phosphates — 

Na 2 HP0 4 + CaCl 2 = CaHP0 4 + 2NaCl. 

2Na 3 P0 4 -f 3 MgHPO, = Mg 3 (P0 4 ) 2 -f- 3 Na 2 HP0 4 . 

The acid phosphates of calcium and magnesium are quite 
soluble in water, imparting the acid reaction. They undergo no 
change by the addition of an acid. With a soluble calcium or 
magnesium salt they change to neutral or basic phosphates — 

CaH 4 (P0 4 ) 2 -f CaCl 2 = 2CaHP0 4 + 2HCI. 
MgH 4 (P0 4 ), + 2MgCl 2 = Mg 3 (POJ 2 + 4HCI. 

Generally there is not a sufficient quantity of calcium and 
magnesium to combine with more than one-third of the phos- 
phoric acid to form basic salts, but when the phosphoric acid 
is in combination with calcium and magnesium, forming acid or 
neutral salts, as is generally the case in normal urine by rendering 
the urine alkaline with a solution of sodium or potassium hydrate, 
all of the calcium and magnesium are precipitated in the form of 
basic salts, and some of the phosphoric acid remains in solution 
combined with the sodium or potassium — 

3CaH 4 (P0 4 ) 2 -J- i2NaOH = Ca 3 'POJ 2 + 4Na 3 P0 4 + I2H 2 0. 

By filtering and adding to the filtrate a solution of calcium 
chloride, basic calcium phosphate precipitates — 

2Na 3 P0 4 + 3CaCl 2 = Ca 3 (POJ 2 + 6NaCl. 
3 



34 CHEMICAL ANALYSTS OF THE URINE. 

As there are nearly two-thirds of the phosphoric acid in normal 
urine combined with the metals of the alkalies, the quantity of 
these phosphates is simply increased by rendering the urine 
alkaline with sodium or potassium hydrate. The neutral phos- 
phates of calcium and magnesium are slightly soluble in water ; 
the latter is more soluble than the former. With acids they 
change to acid phosphates — 

2CaHPO i -f 2HCI = CaH 4 (POJ 2 + CaCl 2 . 

By boiling a saturated solution of either, the solution becomes 
turbid, owing to the formation of some basic phosphate — 
4 MgHP0 4 = Mg 3 (P0 4 ) 2 + MgH 4 (P0 4 ) 2 . 

By this reaction neutral urine becomes cloudy by boiling. The 
basic phosphates of calcium and magnesium are soluble in dilute 
acids, owing to the formation of acid phosphates — 

Ca 3 (POJ 2 + 4HCI = CaH 4 (POJ 2 -f 2CaCl 2 . 

Urine alkaline in reaction, is necessarily turbid, owing to the 
formation of basic phosphates of calcium and magnesium. 

GLYCERIN-PHOSPHORIC ACID. 
Besides the phosphates enumerated above there is a small 
quantity of glycerin-phosphoric acid in normal urine. As its 
name indicates, its component parts are glycerin, C 3 H 5 (OH) 3 , and 

/ OH 

phosphoric acid, PO— oh. The rational formula of glycerin-phos- 
phoric acid is 

/OC 3 H 5 (OH) 2 
PO— OH 
\OH. 

It differs from soluble phosphates in not precipitating with mag- 
nesia mixture (for the preparation of which refer to Chapter viii) 
and ammon. hydrate. Neither is the body precipitated from 
solution when a solution of calcium hydrate and chloride is 
added. By these reagents the phosphoric acid of the phos- 
phates is separated. (Sotnischewsky's method.) Render the urine 
alkaline with the milk of lime, add a solution of calcium chloride 
until a precipitate ceases to form ; filter, and evaporate the nitrate 
to dryness. 

The residue contains the glycerin-phosphoric acid. Treat the 



GLYCERIN-PHOSPHORIC ACID. 35 

residue several times with alcohol, to dissolve out all bodies 
soluble in alcohol, when the residue is dissolved in water and 
treated with a small quantity of magnesia mixture and an excess 
of ammon. hydrate. By this reagent a small quantity of the 
phosphates, that may be in solution, is separated. After standing 
twenty-four hours, filter, and to decompose the glycerin-phosphoric 
acid in the filtrate, render strongly acid with sulphuric acid and 
boil fifteen minutes ; after cooling, render the solution strongly 
alkaline with ammon. hydrate and add some magnesia mixture, 
Crystals of magnesium ammon. phosphate, MgNH 4 P0 4 , will 
separate. To prove that glycerin was in the combination with the 
phosphoric acid evaporate the filtrate to dryness, extract with 
alcohol, and evaporate the alcohol solution of glycerin to dryness, 
when the presence of glycerin in the residue is determined by 
placing some of it in a dry test tube, mixing well with some dry acid 
potassium sulphate, and applying heat. The penetrating odor of 
acrolein is evidence of the presence of glycerin. Moisten a borax 
bead on platinum wire with some of the residue and hold in the 
oxidizing zone of a Bunsen's flame; a green color imparted to the 
flame is characteristic of glycerin. The total quantity of P 2 5 in the 
urine of an adult of twenty-four hours is 2.7 to 3.2 grms., but the 
quantity is subject to vary from 2.5 to 4.5 grms. The relative 
quantity of nitrogen placed at 100 is 18 to 20. From estimations 
of glycerin-phosphoric acid in normal urine, the quantity excre- 
ted in twenty-four hours is 0.030 to 0.060 grm. Occasionally, 
however, no glycerin-phosphoric acid is found in normal urine. 
By the ingestion of soluble phosphates with glycerin (Zuelzer) 
the quantity is increased to 0.354 grm. The quantity is also 
increased, according to Zuelzer, by narcosis of chloroform. The 
total quantity of phosphoric acid in the urine varies according to 
the kind of food ingested. By consulting Table 3 in the Appendix, 
it is found that the relative quantities of nitrogen and P 2 5 , 
in articles of foo'd containing these bodies, differ. In brain, for 
example, the relative quantity of P 2 5 (nitrogen 1 00) is 44, beef 
12.8, etc. Zuelzer found that by the ingestion of different articles 
of food the relative quantity of P 2 5 in the urine varies, as shown 
by Table 4 in the Appendix. 

As brain substance contains lecithin, C 42 H 84 NP0 9 , the quantity 
of P 2 5 excreted by the kidneys is greatly increased by taking 



36 CHEMICAL ANALYSIS OF THE URINE. 

brain as food. The same investigator found that in depressed 
conditions of the nervous system, as during sleep or narcosis 
produced by chloroform, morphine or chloral, the quantity of 
P 2 5 excreted by the kidneys is increased, and on the other hand 
the quantity excreted is diminished by moderate stimulation 
produced by alcohol or strychnin. In fever the relative quantity 
of P 2 5 excreted by the kidneys is less than in health, but during 
convalescence it is greatly increased. 

SULPHUR COMPOUNDS. 
In the urine there are three classes of sulphur compounds: the 
sulphates, as Na 2 S0 4 or CaS0 4 ; the ester compounds of H 2 S0 4 in 
which certain organic radicals take the place of H in H 2 S0 4 , as 
C 8 H 6 N\ S0 ^ and C 6 H 5 \ SQ ^ 

Indoxylsulphuric Acid. Phenylsulphuric Acid. 

and compounds the constitution of which is not fully known. 
These sulphur compounds are separated as they differ in their 
chemical properties. The sulphuric acid of the sulphates is 
separated from the urine by rendering it strongly acid with acetic 
acid, and on adding an excess of a solution of barium chloride, 
BaS0 4 , will separate as a white precipitate. The acetic acid pre- 
vents the precipitation of barium phosphate and has no action on 
the second and third class of compounds, hence they remain in 
solution. The ester compounds of sulphur are decomposed by 
heating with hydrochloric acid, with the liberation of sulphuric 
acid. Separate the BaS0 4 from the solution by filtering, and 
render the filtrate acid with hydrochloric acid, boil a few minutes ; 
a white precipitate, BaS0 4 , forms, there being a sufficient quantity 
of barium chloride in the solution to combine with all of the 
sulphuric acid formed by the decomposition of the ester com- 
pounds. The nitrate of the second precipitation of barium sul- 
phate contains the third class of sulphur compounds, the sulphur 
of which is oxidized to form sulphuric acid. For this purpose, 
concentrate the filtrate by evaporating in a platinum dish on a 
water bath, when the solution is rendered alkaline with a solution 
of sodium carbonate, after which add 7 grms. potassium nitrate 
for every 100 c.c. urine, evaporate to dryness, and heat the residue 
in the dish gradually until it fuses. After cooling, dissolve in 



SULPHUR COMPOUNDS. 37 

water, filter if necessary, render the solution acid with hydro- 
chloric acid and evaporate to a small volume. If the solution is 
still acid transfer to a test tube, and test for sulphuric acid with 
a solution of barium chloride. By this process the sulphur is 
oxidized by the oxygen of the potassium nitrate to form sul- 
phuric acid. The total quantity of sulphur, calculated as H 2 S0 4 , 
excreted by the kidneys in 24 hours is 2 to 4 grms., the relative 
quantity is 18 to 20 nitrogen placed at 100. The total quantity 
varies at different times, depending not only on the quantity of 
albuminous bodies and sulphates in the food, but on the con- 
dition of the liver. It was found by Zuelzer that when the bile 
is drawn off by establishing a fistula, less of the sulphur com- 
pounds are found in the urine. A few hours after a meal, the 
quantity of sulphur compounds in the urine increases, and pre- 
ceding a meal, the quantity decreases. In the febrile state there 
is an increase in quantity, and during convalescence, the liver 
resuming its normal action, less of the sulphur compounds is 
found in the urine. The cause of an increased quantity of the 
sulphur compounds excreted by the kidneys when the liver is 
inactive, is that taurin, C 2 H 7 NS0 3 , of the bile, having sulphur as a 
constituent, is not all absorbed by the blood, as a small quantity 
is found in the faeces. In convalescence the liver is more active, 
an increased quantity of bile passes into the intestines, and much 
of the taurin is excreted with the fseces. During digestion an 
increased quantity of bile is likewise formed, and as a result less 
sulphur is excreted by the kidneys. But as about 80 per cent. 
of the total quantity of sulphur excreted by the kidneys comes 
from albuminous bodies and sulphates of the food, as well as 
from the reduction of tissues, without first entering into the for- 
mation of taurin, slight changes in the physiological conditions 
of the liver bring about no variation in the quantity of sulphur 
compounds in the urine. The quantity of sulphuric acid, in the 
form of phenylsulphuric acid combined with sodium or potassium, 
is increased by the use of carbolic acid, whether used externally 
or internally, so that the urine, after having been rendered strongly 
acid with acetic acid, will yield no precipitate with a solution of 
barium chloride. The phenylsulphates are not poisonous, hence 
the test for sulphates with acetic acid and a solution of barium 
chloride has clinical importance in cases carbolic acid is used for; 



38 CHEMICAL ANALYSIS OF THE URINE. 

as long as the urine forms a precipitate with these reagents, there 
is sufficient H 2 S0 4 to form the phenylsulphates with carbolic 
acid, C 6 H 5 (OH), thereby preventing the toxic effects of the latter. 

CARBONIC ACID. 
Normal urine contains carbonic acid, as acid or normal carbon- 
ates. Acid calcium or magnesium phosphate reacts on the 
carbonates, liberating carbonic acid gas — 

CaH 4 (P0 4 ) 2 -f 2NaHC0 3 = CaIIP0 4 -f Na 2 HP0 4 + 2H 2 + 2C0 2 . 

Urine strongly acid in reaction may contain carbonic acid in 
solution, but not in combination. Normal urine strongly alkaline 
in reaction is alkaline from the presence of the carbonates of 
potassium and sodium. By the process of fermentation ammon. 
carbonate is formed from the decomposition of urea, hence urine 
becomes strongly alkaline. 

Carbonic acid in the urine, if in any considerable quantity, is 
detected by acidifying the urine in a test tube with hydrochloric 
acid, and applying heat, but not to the boiling temperature. 
If effervescence takes place carbonic acid is present — 
NaHC0 3 + HC1 = NaCl + H 2 + C0 2 . 

CALCIUM AND MAGNESIUM COMPOUNDS. 
From the study of the constitution of the phosphates the various 
combinations of calcium and magnesium, as they are found in 
the urine, are understood. To test for calcium and magnesium, 
render the urine distinctly acid with acetic acid, add an excess of 
a saturated solution of ammon. oxalate, the precipitate formed is 
calcium oxalate. Filter, and render the filtrate alkaline with 
ammon. hydrate ; the precipitate formed is magnesium ammon. 
phosphate — 

Mg(C 2 H 3 2 ) 2 2NH 4 C 2 H 3 2 + Na 3 P0 4 = MgNH 4 P0 4 + 3NaC 2 H 3 2 + 

Mag. Ammon. Acet. (double salt). Mag. Ammon. Phos. 

NH 4 C 2 H 3 2 . 

The quantity of calcium, calculated as oxide, excreted by the 
kidneys of an adult in 24 hours is 0.26 to 0.38 grm., and the 
quantity of magnesium, as oxide, is 0.4 to 0.5 grm. The relative 
quantity of calcium oxide — the quantity of nitrogen placed at 1 00 — 
is 0.6 to 1.4, and the relative quantity of magnesium oxide is 0.8 



AMMONIA. 39 

to 1.3. The total quantity of calcium and magnesium oxides 
excreted in a given length of time is subject to variation, but the 
relative quantity of each is more constant. 

By the ingestion of carbonates, or soluble salts of calcium and 
magnesium, the total quantity of each in the urine is increased, 
but the quantity of the salts of each metal absorbed by the blood 
is estimated at only 10 per cent., while 90 per cent, is excreted with 
the faeces. 

In rickets and osteomalacia, the relative quantity of each is not 
increased. 

AMMONIA. 

To test urine for ammonia, to 100 c.c. fresh urine add 200 c.c. 
alcohol; and a solution of platinum chloride acidified with hydro- 
chloric acid. After standing twenty-four hours filter through a 
small filter paper, wash with alcohol, dry and transfer to a dry 
test tube, and heat to redness. The sublimate formed in the test 
tube is ammon. chloride — 

R(NH,) 2 C1 6 = PtCl 4 + 2NH 4 C1. 

Am. Chlor. 

By breaking the test tube and placing the sublimate in a small 
beaker, mixing with some calcium hydrate and moistening with 
water, ammonia gas will be liberated — 

2NH 4 C1 -f Ca(OH) 2 = CaCl 2 + 2H 2 + 2NH a . 

Ammonia gas is known by its penetrating odor and alkaline 
reaction, changing moistened red litmus paper blue, or moistened 
turmeric paper dark red. The quantity of ammonia excreted by 
the kidneys of a person of average weight in twenty-four hours 
is 0.3 to 1.1. The relative quantity — the quantity of nitrogen 
placed at 100 — is 3.5. By the ingestion of any of the mineral acids 
in medicinal doses, no free acid will appear in the urine, but 
the quantity of ammonia is increased. Acids in the blood of 
carnivorous animals increase the quantity of ammonia compounds 
in the urine at the expense of the quantity of urea. The quantity 
of ammonia compounds is also increased by a meat diet and 
diminished by a diet composed principally of vegetables. This 
fact is accounted for on the theory (Schmiedeberg's) of the 
formation of ammon. carbonate in the blood from nitrogenous 
bodies, and by the withdrawal of H 2 from ammonium carbonate 



40 CHEMICAL ANALYSIS OF THE URINE. 

urea is formed, and the ammonia in the urine is a part which has 
been eliminated from the blood before this action took place. 
By the ingestion of ammon. carbonate, the quantity of urea 
excreted is increased, while the amount of ammonia remains 
about the same, but the quantity of ammonia in the urine is 
increased by the ingestion of ammon. chloride. These facts are 
in conformity with the theory. By fermentation of the urine, 
urea is decomoosed, forming ammon. carbonate, and as fermenta- 
tion of the urine may take place in the bladder, the urine in such 
cases is usually strongly alkaline when fresh, the alkalinity being 
due to an increased quantity of ammon. carbonate. To determine 
this condition of the urine, refer to bacteria, page 21. It some- 
times occurs that urine is undergoing the bacterial fermentation, 
and yet has the acid reaction. This is always the case when 
the fermentation is started in acid urine, and insufficient time has 
elapsed to form the amount of ammon. carbonate required to 
neutralize the acid salts in the urine. 

POTASSIUM AND SODIUM. 

Evaporate 50 c.c. urine to dryness in a platinum dish, heat the 
residue over the free flame until the ash appears gray in color. 
When cool, triturate with some water; filter, and into the filtrate 
dip a piece of platinum wire which has been heated to redness 
some time. Now place the end of the wire in the colorless flame 
(Bunsen's), when the sodium present will impart a yellow color 
to the flame. 

The presence of potassium may be determined by looking at 
the yellow sodium flame through a piece of glass colored blue by 
cobalt, when the yellow rays are arrested and a violet color is 
perceived, due to the presence of potassium. Not having cobalt 
glass at hand, concentrate the solution (filtrate) to one or two 
cubic centimetres, and to this add a strong solution of tartaric 
acid ; a precipitate will form — 

KC1 + C 4 H 6 6 = C 4 H 5 K0 6 + HC1. 

Acid Potass. Tart. 

After standing several hours, filter, wash, dry, and employ the 
color test, as above, by moistening the end of the wire with some 
dilute hydrochloric acid, and dipping into the dry powder. In 
normal urine there is more sodium than potassium. The total 



POTASSIUM AND SODIUM. 41 

quantity of sodium excreted by the kidneys of an adult in 
twenty-four hours is 4 to 5 grms. and of potassium 2 to 3 grms. 
The relative quantity of sodium — nitrogen 100 — is 35 to 40, and 
of potassium 25. Potassium is a constituent of many of the 
tissues of the body, while sodium compounds are held in solution 
by the fluids of the body. It was found by Zuelzer that for 100 
parts nitrogen in the brain there are 21 parts potassium and 8.7 
parts sodium, and for the same number of parts nitrogen in 
muscular tissue there are 6.1 parts potassium and 1.1 part sodium. 
As a result of there being more potassium than sodium in the 
tissue, when there is rapid desassimilation, as in fevers, the 
quantity of potassium excreted by the kidneys exceeds that of 
sodium. 



CHAPTER III. 

Bodies in Diseased Urine — Albuminous Bodies, including Mucine — Serum Albu- 
men — Globuline — Hemialbumose — Peptone — Mucine — Color Tests for Albuminous 
Bodies — Millon's Reagent — Table of Reactions of Albuminous Bodies, including 
Mucine — Table of Color Tests — Methods for Determining the Presence of Albu- 
minous Bodies — The Nitric Acid Test — The Acetic Acid and Sodium Chloride 
Test — Determination of the Presence of Hemialbumose, Peptone. Globuline, and 
Mucine in Aluminous Urine — Diabetic Sugar — Tests for Sugar in Urine — Trom- 
mer's Test Modified by Salkowsky — Test with Fehling's Solution — The Ferment- 
ation Test — The Phenylhydrazin Test — Methods for Separating Small Quantities 
of Sugar from Urine — Inosit — Test for Inosit. 



ALBUMINOUS BODIES. 
The urine may be regarded as a filtrate of the blood, and by 
certain diseases of the kidneys or changes in the constitution of 
the blood or variation in pressure of the blood in the capillaries 
of the kidneys, the various albuminous substances of the blood 
plasma appear in the urine ; and while in the majority of cases it 
is sufficient for the physician to know that "albumen" is present, 
yet in some cases it is doubtless important to determine the pres- 
ence of other members of this group of bodies. What is gener- 
ally recognized as albumen in the urine is a mixture of serum 
albumen and globuline, as these bodies are precipitated by the 
reagents usually employed in testing for albumen. The quantity 
of albumen excreted in twenty-four hours differs greatly ; 7 grms. 
may be considered the average quantity, although as much as 
30 grms. are sometimes excreted. The principal albuminous body 
found in the urine is serum albumen ; the other members of the 
class are found in small quantities, and in many cases are absent. 

SERUM ALBUMEN. 

1. Alcohol precipitates albumen from aqueous solutions, hence, 
by the addition of alcohol to albuminous urine, the albumen 
separates in flake-like bodies. 

2. Nitric acid precipitates albumen from the urine when added 
in considerable quantity; in great excess, however, it redissolves. 
To one volume albuminous urine add six vols, water, when small 
quantities of nitric acid are added, shaking after each addition. 

42 



PEPTONE. 43 

Albumen will precipitate and redissolve until the acid is added in 
certain quantity, when the albumen becomes insoluble; therefore 
nitric acid in small quantities does not precipitate albumen. The 
object of adding water to the urine is to dilute the solution of 
neutral salts of the urine, as they aid small quantities of nitric 
acid in precipitating albumen unless in dilute solutions. 

3. Albumen in the urine precipitates by boiling, if the urine is 
strongly acid in reaction, but by the addition of a solution of 
sodium hydrate or carbonate, the urine will remain clear by boil- 
ing. By heating urine, neutral or slightly acid in reaction, basic 
phosphates of the metals of the alkaline earths precipitate (refer 
to phosphates in the urine). 

4. Acetic acid does not precipitate albumen in the urine, but 
when a strong solution of common salt, sodium or magnesium 
sulphate is added to the urine, acidified with acetic acid, and the 
urine is boiled, the albumen will precipitate; but without the addi- 
tion of a solution of one of these salts, an excess of acetic acid is 
avoided or some of the albumen may remain in solution. 

5. By rendering albuminous urine strongly acid with acetic 
acid, and adding a few drops of a solution of potassium ferro- 
Cyanide, albumen will precipitate without the application of heat. 

GLOBULINE AND HEMIALBUMOSE. 

These bodies are similar in many of their properties. Both 
are insoluble in water and alcohol, but in water holding in solu- 
tion neutral salts, alkalies or acid salts, as the urine, they are 
soluble. Globuline separates from the urine by heating to the 
boiling temperature with a moderate excess of nitric acid, while 
hemialbumose remains in solution, but separates as the urine 
cools. Hemialbumose remains in solution when the urine is 
treated with y& its volume of a saturated solution of common 
salt, and acidified with acetic acid, boiled, and filtered while hot. 
By this process serum albumen, globuline, and mucine are sepa- 
rated from solution. 

PEPTONE. 

Peptone is soluble in water. Peptone, serum albumen, and 
hemialbumose are precipitated from solution in the urine by tannic 
acid, corrosive sublimate and phosphortungstate of sodium with 
hydrochloric acid. Peptone is not precipitated from solution in 



44 CHEMICAL ANALYSIS OF THE URINE. 

the urine by nitric acid or acetic acid with a strong solution of 
common salt or acetic acid and potassium ferrocyanide. 

MUCINE. 

Although mucine is a constituent of normal urine, yet it is often 
found in greatly increased quantities from cystitis. It is sometimes 
in shreds or flakes, usually, however, it is in solution. Mucine is 
precipitated from solution in the urine by adding two volumes of 
alcohol. It is also precipitated by a solution of tannic acid, 
acetic acid or a solution of neutral lead acetate. It is quite 
soluble in water rendered alkaline with potassium or sodium 
carbonate. 

COLOR TESTS FOR ALBUMINOUS BODIES. 

Wash some coagulated albumen, saturated with a dilute solution 
of copper sulphate, on a filter, then transfer some of the coagula 
to a test tube and treat with a solution of sodium hydrate. The 
solution becomes deep blue in color. 

A solution of peptone rendered alkaline with sodium hydrate, 
and treated with a small quantity of a solution of copper sulphate, 
yields a violet color. A solution of hemialbumose, with these 
reagents, yields a purple violet color. 

MILLON'S REAGENT. 
For the preparation of Millon's reagent refer to Chapter vin. To 
four or six volumes of water in a test tube, containing a small 
coagula of albumen, one volume of Millon's reagent is added 
and the mixture heated to the boiling temperature. Particles 
of albumen will float on the surface of the fluid, if there is an 
excess of the reagent ; if not, they will settle ; in either case they 
are colored light red. Reactions of albuminous bodies with 
reagents, generally employed for their detection in the urine, are 
shown by the following table : — 



REACTIONS OF ALBUMINOUS BODIES IN THE URINE. 45 

REACTIONS OF ALBUMINOUS BODIES IN THE URINE. 



REAGENT. 


SERUM GLOBULINE. HEMIALBUMOSE. 
ALBUMEN. 


PEPTONE. MUCINE. 


To the urine add nitric ppt. when cold, 
acid in excess ; the ! 1 if the acid is 
urine is then heated to not in great 




ppt., and by 
continued 
boiling de- 
composes. 


ture. 




redissolves 
when heated, 
imparting a 
yellow color. 




To 5 vols, urine add 
i vol. saturated solu- 
tion sodium chloride, 
render strongly acid 
with acetic acid and 
heat to the boiling tem- 
perature. 


ppt. 


ppt. 


No ppt., but 
when the urine 
becomes cold, 
a ppt. forms. 
If the urine 
be saturated 
with sodium 
chloride, the 
hemialbumose 
will precipi- 
tate and not 

redissolve 
when the urine 
is heited. 




ppt. 




To i vol. urine add 3 
vols, alcohol, 90 per 

cent., when the urine : ppt. ppt. ppt. 
is rendered acid with 
acetic acid. 


ppt., partly. 


ppt. 


To the urine add a sol. 
of phosphortungstate 
of sodium, acidified 
with acetic acid, or a 
solution of phosphor- 
tungstic acid. 


ppt. ppt. ppt. 


ppt. 


ppt., if the urine 
is decidedly 
acid with 
acetic acid, 
otherwise no 
ppt. forms. 


1 
To the urine add acetic 
acid in excess, when a 1 


ppt., but redis- 
solves if the 




ppt. by the ace- 
tic acid. 


of potassium ferrocya- 
nide is added. 






to near the 
boiling tem- 
perature. 




To the urine add a 
strong solution of tan- 
nic acid. 


ppt 


ppt. 


ppt. 


ppt. 


ppt., partly 
from urine, if 
neutral. The 
ppt is in- 
creased in 
quantity by 
adding a solu- 
tion of sodium 
chloride. 


To the urine add acetic 






PPt. 


the application of hear . 







46 CHEMICAL ANALYSIS OF THE URINE. 

COLOR TESTS. 
ALBUMINOUS BODIES, HAVING BEEN SEPARATED FROM THE URINE. 



REAGENT. 


SERUM 
ALBUMEN. 


GLOBULINE. 


HEMIAI.BUMOSE. 


PEPTONE. 


MUCINE. 


Having saturated the 
coagulae with a solu- 
tion of copper sul- 
phate, introduce into 
a test tube, and boil 
with an excess of a 
solution of sodium hy- 
drate. 


blue color. 


blue color. 


purple violet or 

blue with more 

copper 

sulphate. 


purple or 
violet color. 


blue color. 


Millon's Reagent — em- 
ploy as above, page 44. 


red color. 


red color. 


red color. 


red color. 


red color. 



METHODS FOR DETERMINING THE PRESENCE OF ALBUMINOUS BODIES IN 

THE URINE. 

If the urine is turbid, filter, and test the nitrate. In no 
case should cloudy or turbid urine be employed in testing for 
albumen. In case the filtrate is turbid, treat the urine with a 
solution of magnesium sulphate, add a small quantity of a 
solution of sodium carbonate, shake well and filter; the filtrate 
will be clear. 

1. THE NITRJC ACID TEST. 

To the urine in a test tube add J^ its volume of nitric acid, 
specific gravity 1.2; warm a short time, but not to the boiling 
point. If a flocculent precipitate forms, it is albumen (serum 
albumen and globuline). By transmitted light the fluid in the 
test tube will appear clear. 

The conditions to be observed in the employment of this test 
are : The urine should be clear, the acid of certain strength 
(sp. gr. 1.2), added in definite quantity, and the urine should not 
be boiled. 

2. ACETIC ACID AND SODIUM CHLORIDE TEST. 

Render the urine strongly acid with acetic acid, and to five 
volumes urine add one volume of a saturated solution of common 
salt,* and heat to the boiling point. If a precipitate forms, it is 

* The saturated solution of common salt is prepared by dissolving 350 grms. sodium 
chloride in one litre water: filter the solution, if it is not clear. 



ACETIC ACID AND SODIUM CHLORIDE TEST. 47 

composed of serum albumen and globuline while the solution is 
hot, but as it cools, hemialbumose also separates. The conditions 
for the employment of this test are : The solution of sodium 
chloride should be saturated and added in definite quantity, and 
the urine should be strongly acid with acetic acid. 

METHOD FOR DETERMINING THE PRESENCE OF HEMIALBUMOSE IN 
ALBUMINOUS URINE. 

To five volumes urine add one volume saturated solution of 
common salt, and render strongly acid with acetic acid; heat to 
the boiling point and filter while hot. If hemialbumose is present, 
the filtrate will become cloudy as it cools. To prove that the 
precipitate so formed is hemialbumose, to the filtrate add an equal 
volume of the saturated solution of sodium chloride. Hemialbu- 
mose will precipitate. Filter, and dissolve the precipitate in strong 
nitric acid; warm gently; the acid solution becomes deep yellow 
in color if the body is hemialbumose. 

METHOD FOR DETERMINING THE PRESENCE OF PEPTONE IN ALBUMINOUS 

URINE. 

To separate albuminous bodies, except peptone, from the urine, 
5CO c.c. urine (Hofmeister) is treated with 10 cc concentrated 
solution of sodium acetate, when a solution of ferric chloride 
is added until, after mixing, it assumes a red color. Boil the 
solution and filter while hot. The filtrate should yield no pre- 
cipitate with acetic acid and potassium ferrocyanide. Peptone is 
separated from the filtrate by adding an excess of a solution of 
tannic acid, and, after standing twenty-four hours, filtering. Wash 
the precipitate with water containing some tannic acid and mag- 
nesium sulphate. To separate the tannic acid from the peptone, 
the precipitate is triturated with a saturated solution of barium 
hydrate in a small mortar, while some crystallized barium hydrate 
is added. The mixture is then introduced into a small beaker 
or dish, and heated on a sand bath to the boiling point. After 
standing a few minutes, filter, and test the filtrate for peptone, 
employing the color tests as above. 

METHOD FOR DETERMINING THE PRESENCE OF GLOBULINE AND SERUM 
ALBUMEN IN ALBUMINOUS URINE. 

Filter the urine if necessary, and if the filtrate is acid or alka- 
line, render it neutral by employing a solution of sodium car- 



4S CHEMICAL ANALYSIS OF THE URINE. 

bonate or dilute acetic acid. Shake or stir the urine with pul- 
verized magnesium sulphate until it ceases to dissolve. Globuline 
will separate as a precipitate. Filter, and wash the precipitate 
with a saturated aqueous solution of magnesium sulphate until 
the wash water ceases to form a precipitate when acidified with 
acetic acid and heated to the boiling point. Transfer the pre- 
cipitate to a beaker and add water until it dissolves. Heat the 
solution to the boiling point, when, if globuline is present, it will 
precipitate. The filtrate, after treating with magnesium sulphate, 
contains the serum albumen, to precipitate which render the 
solution strongly acid with acetic acid and heat to the boiling 
point. In the separation of globuline from albuminous urine, 
instead of employing magnesium sulphate, ammon. sulphate may 
be employed (Pohl's method). For this purpose render the urine 
slightly alkaline with ammon. hydrate, and, having stood several 
hours, filter, to separate the phosphates of calcium and magne- 
sium. To one volume of the filtrate add one volume of a satu- 
rated solution of ammon. sulphate. If a precipitate forms, it is 
globuline. 

METHOD FOR DETERMINING THE PRESENCE "OF MUC1NE IN ALBUMINOUS 

URINE. 

To one volume of urine add three volumes strong alcohol, 
stir, and let stand several hours. Mucine and all albuminous 
bodies precipitate. Filter and wash the precipitate with alcohol. 
Treat the precipitate on the filter with warm water. The filtrate 
contains the mucine, to precipitate which render strongly acid 
with acetic acid ; if the solution becomes turbid, mucine is 
present. Refer to reactions of albuminous bodies, including 
mucine, page 45. In urine not containing albuminous bodies, 
the presence of mucine is determined by diluting one volume 
of urine with one volume water and adding an excess of acetic 
acid ; mucine precipitates, causing the urine to appear cloudy 
or opaque. The object of diluting the urine is to overcome the 
tendency of the neutral salts of the urine to dissolve the mucine 
in the presence of an excess of acetic acid. 



DIABETIC SUGAR. 49 

DIABETIC SUGAR. 
Glucose, grape sugar, corn sugar, and starch sugar are synony- 
mous with diabetic sugar. Diabetic sugar is a crystalline body 
containing water of crystallization. The formula is — 

C 6 H 12 6 , H 2 0. 

By drying in a desiccator over concentrated sulphuric acid, 
it loses its water of crystallization when it melts at 146 . 
Diabetic sugar is soluble in water, somewhat soluble in alcohol 
and insoluble in ether. It is not easily decomposed by acids, 
but decomposes when heated in solution with potassium or 
sodium hydrate. With potassium hydrate, sodium chloride, or 
lead acetate with ammon. hydrate, it separates as an insoluble 
body. 

Diabetic sugar has the chemical property of an aldehyde, hence 
it is easily oxidized to form an acid. 

C 6 H 12 6 + 2 = H 2 + C 6 H 10 O 7 . 

It is on this property that depends one of the most important 
tests for it in the urine. By the action of the yeast ferment, it 
decomposes into alcohol and carbonic acid — 

C 6 H 12 6 = 2C 2 H 6 + 2C0 2 . 

Diabetes Mellitus is a disease characterized by sugar in the 
urine; but it is more than this; one of its prominent features 
is, that there is rapid desassimilation or waste of the tissues 
in acute cases, hence there is more urea excreted in a given 
length of time than could be formed without loss in weight, 
taking into account the quantity of nitrogenous food consumed. 
With a meat or nitrogenous diet a greater amount of sugar 
appears in the urine than could be formed from the starchy or 
saccharine elements of the food; consequently, in the reduction of 
albuminous compounds in the blood, sugar is probably one of 
the products. There are increased quantities of phosphoric and 
sulphuric acids excreted by the kidneys in diabetes, and the 
quantity of water drained from the blood is likewise increased, 
often amounting to four times the normal quantity. Diabetic 
urine is generally light colored, high specific gravity, caused by 
the sugar it contains, and acid in reaction. 
4 



50 CHEMICAL ANALYSIS OF THE URINE. 

TESTS FOR SUGAR IN THE URINE. 
TROMMER'S TEST MODIFIED BY SALKOWSKY. 

For the employment of Salkowsky's modified form of Trom- 
mer's test, solutions of sodium hydrate and copper sulphate of 
known strength are used. 

ioo grms. sodium hydrate are dissolved in 300 cc. water, 
and after standing a week, if a sediment forms, decant the 
solution from it. This solution should be kept in a glass- 
stoppered bottle, but care should be taken that the stopper 
and ground surface of the neck of the bottle be free of the 
alkaline solution, or difficulty in removing the stopper may be 
encountered. Dissolve 30 grms. crystallized copper sulphate in 
300 cc. water. To three volumes of urine add one volume of the 
solution of sodium hydrate, and of the mixture fill a test tube 
one-half full and add the solution of copper sulphate, drop by 
drop, shaking the fluid after each, addition, until the copper 
hydrate ceases to dissolve. Heat the mixture to near the boiling 
point. If sugar is present, yellow or red cuprous hydrate or 
oxide will separate. The reduction usually takes place first in 
the upper stratum of the fluid. The mixture is not boiled, as a 
few degrees below the boiling point are sufficient for the reduction 
to take place. There is no difficulty of determining sugar in the 
urine in the majority of cases, especially if the sugar is in consid- 
erable quantity, but as uric acid, coloring matters, kreatinin and 
other bodies the constitution of which is not known, reduce 
copper oxide to a certain extent, it sometimes becomes a question, 
in the employment of this test, if sugar is in the urine. 

FEHLING'S SOLUTION. 

Qualitative tests for sugar in the urine are made with Fehling's 
solution. This solution is prepared by introducing 17.32 grms. 
crystallized copper sulphate in a 250 cc. graduated flask, when 
water is added, and, the salt having dissolved, the flask is filled 
with water to the mark and the solution is well mixed by shak- 
ing. Introduce into a 250 cc graduated flask 86.5 grms. potas- 
sium sodium tartrate and 25 grms. sodium hydrate; add water, and 
when solution has taken place fill with water to the mark, and 
finally mix well by shaking. An equal volume of each solution, 
well mixed, is Fehling's solution. In preparing the solutions for 
qualitative tests, the weights and measurements may be approxi- 



TESTS FOR SUGAR IN THE URINE. 51 

mative. It is not practical to keep Fehling's solution on hand a 
great length of time, as it is liable to decompose. 

Into a small beaker introduce about 10 cc. Fehling's solution 
and 50 cc. water, heat on a sand bath to near the boiling temper- 
ature, when 10 cc. urine is introduced from a pipette, the temper- 
ature of the solution still kept near the boiling point a few 
moments. If the color of the solution becomes green, continue 
the addition of urine in quantities of 10 cc. until 40 cc. is added. 
The beaker is then placed in some cold water, so that in case 
cuprous oxide is formed it may settle rapidly. When cold the 
solution is decanted, and if cuprous oxide has formed it may be 
seen on the bottom and sides of the beaker, or, if the cuprous 
oxide is in small quantity, by surrounding the end of a stirring 
rod with some filter paper and wiping the surfaces of the beaker, 
the red oxide is seen on the paper. If the urine contains a 
decided quantity of sugar, reduction of copper oxide takes place 
at once, and the solution becomes brick-dust red in color from the 
presence of cuprous oxide. In the employment of this test, 
Fehling's solution diluted with water, as above, should be heated 
in a beaker to near the boiling point for a short time, and, when 
cold, examined, to see if any reduction of copper oxide has taken 
place. Without making this preliminary test, mistakes are liable 
to occur. Occasionally urine, when heated with Fehling's solution, 
diluted as above, w r ill yield a light yellow precipitate, cuprous 
hydrate, from the presence of reducible substances other than 
sugar, but when this takes place the precipitate usually forms 
while the solution is cooling. In cases of doubt, the urine is 
decolorized by one of the following methods : — 

1. To the urine add a saturated solution of neutral and basic 
lead acetate until, after stirring and settling, a precipitate ceases to 
form. An excess of the lead salts is avoided. Filter; the filtrate 
will be nearly colorless. 

2. A saturated solution of mercuric chloride — corrosive subli- 
mate — is added to the urine until a precipitate ceases to form. 
The mixture is then rendered alkaline with a solution of sodium 
carbonate, when it is filtered. The filtrate obtained by the em- 
ployment of either process, 1 or 2, is examined for sugar by 
Salkowsky's form of Trommer's test or by the employment of 
Fehling's solution. 




52 CHEMICAL ANALYSIS OF THE URINE. 

Coloring matters of the urine are removed by boiling the urine 
with pulverized animal charcoal, but either of the methods de- 
scribed above is preferable, as sugar is absorbed by the charcoal. 

THE FERMENTATION TEST. 
For this test a piece of combustion tube about 20 cm. long is 
sealed at one end, and near the other end it is drawn out some- 
what and bent, as shown in Fig. 2. The capacity of the tube to 
Fig the part bent is about 20 cc. Before 

mixing the urine with the yeast, boil it 
several minutes so as to drive off C0 2 
and other gases. If the urine is alka- 
line, it is rendered distinctly acid with 
a solution of tartaric acid before boiling. 
Pressed yeast is mixed with a small 
quantity of water, filtered and washed 
with water on the filter several times, 
that saccharine bodies which might be 
present may be removed. About 30 cc. of the urine, having been 
boiled and cooled, is introduced into a small mortar and triturated 
or mixed with about 0.5 grm. of the washed yeast by means of a 
stirring rod. If the mixture is but faintly acid in reaction, it is 
rendered distinctly acid with a solution of tartaric acid, carefully 
avoiding the addition of more than is necessary. The tube is then 
filled with the mixture and placed in position, when a small 
quantity of mercury is introduced into the open end of the tube 
to prevent communication between the urine in the different parts 
of the tube. The apparatus is kept between 15 and 25 ° C. for 
twenty-four hours. If sugar is present, C0 2 gas will collect in 
the sealed extremity of the tube within six hours. In the 
employment of this test it is necessary to determine the relative 
quantity of gas evolved from normal urine. For this purpose a 
test is made by triturating about the same quantity of washed yeast 
with normal urine, having been acidified, if necessary, boiled and 
subjected to the same temperature as that of the urine supposed 
to contain sugar. Normal urine, by the yeast fermentation, yields 
a small quantity of gas, but the amount is greatly diminished by 
boiling the urine. If the suspected urine yields a greater volume 
of gas, sugar is present. If no gas is evolved in either, the yeast 



PHENYLHYDRAZIN TEST. 53 

may not be effective; it is then triturated with some normal urine, 
acidified, if necessary, with tartaric acid, and to the mixture a 
small quantity of cane sugar is added, and the mixture is then 
introduced into the tube and subjected to a favorable tempera- 
ture, 20° C. 

PHENYLHYDRAZIN TEST. 

The employment of this reagent for the detection of sugar in 
urine has been used to a very limited extent, as its use for this 
purpose is of recent discovery. 50 cc. urine, supposed to con- 
tain sugar, is introduced into a beaker, when 2 grms. phenylhy- 
drazin hydrochlorate with 1.5 grm. sodium acetate, or 1 grm. of 
the latter if the urine is not decidedly acid, is added. Unless the 
urine is nearly colorless, add 20 cc. water. The beaker is then 
placed in a water bath and warmed gently one hour. If sugar is 
present crystals of phenylglucosazon will form. It was at first 
supposed that the form of crystals of this body, as found by the 
microscope, would be sufficient for determining the body ; but it 
was subsequently found that the only positive evidence that the 
body is phenylglucosazon is obtained by separating it from the 
urine by filtering, washing with a small quantity of water and 
dissolving in a small quantity of dilute alcohol, the body crystal- 
lizes out by evaporating at a low temperature. 

This process is repeated two or three times, when the crystals are 
collected, dried in a desiccator over concentrated sulphuric acid and 
the temperature at which the body melts determined. Phenylglu- 
cosazon melts at 204 to 205 ° C. For the purpose of determining 
the melting point, draw out a piece of thin glass tubing in a Bun- 
sen's flame or a spirit lamp, so that the sealed capillary extremity 
is 2 or 3 cm. from where the tube is of original diameter. The 
tube is broken, by means of a file, near where the contraction 
begins, and a small quantity of the dry body is introduced into 
the sealed extremity. The piece of tubing is now attached to 
a thermometer by means of a small rubber band (obtained by 
making a section of a rubber tube). The capillary end of the 
tube containing the body is placed adjacent to the bulb of the 
thermometer, and the bulb with the tube is introduced into con- 
centrated sulphuric acid in a beaker, when the acid is heated 
gradually. As the mercury ascends to 204 the substance will 
begin to show evidence of fusion, providing the increase in tern- 



54 CHEMICAL ANALYSIS OF THE URINE. 

perature is gradual and the heat be equally diffused by stirring 
the sulphuric acid with a glass rod. 

METHODS FOR THE SEPARATION OF SUGAR ^ROM THE URINE. 

In case urine contains sugar in small quantity so that the 
results of the tests for it are not satisfactory, it may be separated 
from the urine by either of the following methods, and the 
aqueous solution tested for sugar : — 

I. About one litre urine, if not acid in reaction, is rendered 

acid with hydrochloric acid and evaporated on a water bath to 

near dryness, when the residue is treated two or three times with 

small quantities of strong alcohol, and the alcohol solution is 

filtered. To the filtrate add an alcohol solution of potassium 

hydrate, stir and let stand two hours. If sugar is present a 

precipitate will form — 

(C 6 H 12 6 ) 2 K 2 0, 

which is separated from the alcohol solution by decantation and 
washed once with strong alcohol. Dissolve the precipitate with 
water, and neutralize the solution with dilute acetic acid; treat the 
neutral solution with a solution of lead acetate until it ceases to 
form a precipitate, when it is filtered and the lead is separated 
from the filtrate with sulphuretted hydrogen gas, H 2 S. The solu- 
tion is filtered from the PbS and the filtrate is concentrated by 
evaporation on a water bath. During the evaporation the H 2 S 
will escape. Instead of separating the lead by means of H 2 S, a 
solution of pure oxalic acid may be employed, and the excess of 
oxalic acid removed from the filtrate by mixing well with pure 
amorphic calcium carbonate. Filter ; concentrate the filtrate by 
evaporating and test the concentrated fluid by any one of the 
tests above given. 

BRUECKE'S METHOD. 

To one litre fresh urine add a solution of lead acetate until a 
precipitate ceases to form ; filter, and treat the filtrate with one or 
two grms. finely powdered basic lead acetate, and after mixing 
well with a stirring rod, add ammon. hydrate in quantity that 
after stirring well the odor of the gas is perceptible. The sugar 
combines with the lead hydrate, as it is precipitated with ammon. 
hydrate. Filter, and wash once with water, and transfer from the 
filter to a beaker by means of a fine stream of water from a wash 



IN0S1T. 55 

bottle. The ammonia gas is driven off by heating to near the 
boiling point, until the vapors given off are no longer alkaline in 
reaction, when sulphuretted hydrogen gas is passed through the 
mixture, sugar is set free and PbS formed. Filter, and concentrate 
the filtrate by evaporating on a water bath. The fluid so concen- 
trated, and free of H 2 S gas, is tested for sugar by any of the tests 
given above. In the employment of this method, the part of the 
process requiring special attention is the complete separation of 
ammonia by heating the mixture; for, if some remains in solution, 
ammon. sulphide will form when the solution is treated with H 2 S, 
which would form CuS with CuO, if either Trommer's test or 
Fehling's solution be employed for the detection of sugar. 

Albumen in urine is first removed before the urine is tested 
for sugar. For this purpose a solution of tannic acid may be 
employed. Having precipitated the albumen, it is separated by 
filtering and the filtrate is tested for sugar. When possible, fresh 
urine is employed for the detection of sugar. 

INOSIT. 
Inosit is isomeric with diabetic sugar. It is a crystalline body, 
soluble in water, but insoluble in absolute alcohol and ether. 
It differs from diabetic sugar in forming an insoluble compound 
in neutral solutions with basic lead acetate ; and with yeast it 
does not change into alcohol and carbonic acid, neither does it 
reduce copper oxide. 

SCHERER'S TEST FOR INOSIT. 

When an aqueous solution of inosit and calcium chloride is 
evaporated to dryness and the residue treated with ammon. 
hydrate, a rose-red color is produced. Inosit sometimes appears 
in albuminous and sometimes in diabetic urine. Inosituria is 
a disease dependent on other diseases, although in some cases 
diabetes mellitus changes into inosituria, that is, sugar disappears 
from the urine and in its place inosit appears. 

TEST FOR INOSIT IN URINE. 

One litre of urine is rendered slightly acid, if not already so, 
with hydrochloric acid and treated with a solution of neutral lead 
acetate, until a precipitate ceases to form, avoiding the addition 



56 CHEMICAL ANALYSIS OF THE URINE. 

of more than is necessary. Filter, and concentrate the filtrate to 
one-fourth its volume by evaporating on a water bath, when the 
solution is treated, while still warm, with basic lead acetate, until 
a precipitate ceases to form. After standing twenty-four hours, 
filter, transfer the precipitate from the filter to a beaker by means 
of a fine stream of water from a wash bottle, and through the 
mixture pass sulphuretted hydrogen gas. Separate the PbS by 
filtering after standing a few hours ; if uric acid has separated, 
decant the fluid into an evaporating dish, evaporate to the con- 
sistence of syrup and triturate with absolute alcohol. The residue 
or precipitate, having been washed with absolute alcohol, is dis- 
solved in hot water and tested for inosit, employing Scherer's 
test. To separate inosit from aqueous solution, add four volumes 
strong alcohol and sufficient ether to produce a cloudiness, and 
by standing inosit will crystallize. 



CHAPTER IV. 

Bodies in Diseased Urine, Continued — Biliary Acids — Tests for Biliary Acids in 
Urine — Biliary Coloring Matters — Tests for Biliary Coloring Matters — Coloring 
Matters of the Blood — The Spectroscope — Spectroscopic Test for Haemoglobin, 
Oxyhemoglobin and Methaemoglobin — Heller's and Struve's Tests for Coloring 
Matters of the Blood in the Urine — The Elements of Blood in Urine — Blood Cor- 
puscles — Separation of Fibrin from Urine — Tests for Fibrin — Leucin — Separation 
of Leucin from Urine — Tyrosin — Separation of Tyrosin from Urine — Fat — Tests 
for Fat in Urine — Separation of Fat from Urine — Dreschel's Apparatus — Choles- 
terin and Lecithin — Method for Separating Cholesterin from Urine — Tests for 
Neurin and Glycerin- Phosphoric Acid Produced from the Action of Barium 
Hydrate on Lecithin. 

BILIARY ACIDS. 
By certain pathological processes in the liver, the bile is 
retained and absorbed or the liver fails to eliminate the constit- 
uents of bile absorbed from the intestines, when the biliary acids 
and coloring matter with its oxidation products appear in the 
urine. That the biliary acids are ever found in urine was long a 
question, until reactions of them became known, by which their 
presence is determined. Biliary acids combined with sodium 
differ in constitution in the bile of different animals. In the bile 
of the ox there are two acids, glycocholic (C 26 H 43 N0 6 ) and tauro- 
cholic (C 26 H 45 NS0 7 ) acids. Each of these acids absorbs water 
and yields as product of decomposition cholic acid, C 2 4H 40 O 5 . 
Besides this body, glycocholic acid yields glycocol, C2H5NO2, and 
taurocholic acid, taurin, C 2 H 7 NS0 3 . The composition and phys- 
ical properties of the acids in human bile are unknown, but the 
acid product corresponding to cholic acid is anthropocholic acid, 
QsH^O^ and in properties it is similar to cholic acid, as it is 
formed by boiling a solution of human bile with barium hydrate. 
It is monobasic, crystalline, and insoluble in water, but soluble 
in alcohol and ether. The acids in human bile corresponding to 
glycocholic and taurocholic acids have not been separated. 

PETTENKOFFER'S TEST FOR BILIARY ACIDS. 

To three parts of a solution of biliary acid compounds add 
two parts of strong sulphuric acid gradually, so that the tempera- 
ture remains under 6o° C, then add three drops of a solution of 

57 



58 CHEMICAL ANALYSIS OF THE URINE. 

cane sugar (i part cane sugar and 4 parts water) and shake well. 
The solution becomes violet in color. 

TEST FOR BILIARY ACIDS IN URINE. 

On account of the presence of indican in the urine, the pres- 
ence of biliary acids in icteric urine cannot always be determined 
without first separating them from the urine. As the biliary 
acids are never found in the urine in great quantity, one litre of 
the urine (Neukomm) is evaporated on a water bath to near dry- 
ness, the residue triturated with strong alcohol, filtered and the pro- 
cess repeated several times. The alcohol filtrate is evaporated to 
dryness on a water bath, treated with absolute alcohol several 
times, and the alcohol solution is filtered. The biliary acids are 
thus separated from bodies in the urine insoluble in alcohol. Evap- 
orate the alcohol filtrate on a water bath to dryness, and treat the 
residue with water, and after stirring several minutes add finely pul- 
verized basic lead acetate until a precipitate ceases to form, when a 
few drops of a solution of the salt is added. After standing several 
hours the solution is filtered and the precipitate washed with 
some water. Transfer the precipitate to a flask, treat with alcohol, 
85 per cent., boil on a water bath a short time, and filter while 
hot. The lead compounds of the biliary acids are insoluble in water 
but soluble in alcohol while hot. To the filtrate add a solution 
of sodium carbonate to alkaline reaction and evaporate on a 
water bath to dryness. Boil the residue with absolute alcohol, 
filter while hot into a flask, and, having become cold, add ether 
until a precipitate forms, close the flask with a stopper, and let 
stand twenty-four hours. Sodium salts of the biliary acids will 
crystallize. Decant the fluid and dissolve the crystals in some 
water, and test the solution with PettenkofTer's reagents. 

COLORING MATTERS OF THE BILE IN URINE. 
Bilirubin, C 16 H 18 N 2 3 , is the principal coloring matter of the 
bile. Of this, however, there are several oxidation products, the 
principal one being biliverdin (C 16 H2oN 2 O g , according to Stsedeler). 
Bilirubin is a solid, insoluble in water but soluble in chloroform. 
Solutions of alkaline carbonates or hydrates dissolve it. With 
calcium salts it forms insoluble bodies in water. Bilirubin, when 
dissolved in an aqueous solution of sodium hydrate or carbonate 



TESTS FOR COLORING MATTERS IN URINE. 59 

and exposed to the air, oxidizes to form biliverdin. Biliverdin is 
a green solid, insoluble in water and chloroform, but soluble in 
alcohol, and like bilirubin, it dissolves in solutions of the alkaline 
carbonates or hydrates, and precipitates from solution with sol- 
uble salts of calcium. Bilirubin, absorbed by the blood from the 
intestines, is separated from the blood by the liver, and not by the 
kidneys, unless the quantity in the blood is greatly increased. 
Pathologists have found that bilirubin is also formed in the blood 
by processes which diminish the number of red corpuscles, and 
that the liver, although not structurally diseased, may fail to 
take from the blood the amount of bilirubin formed, in which 
event the kidneys accomplish the work. This form of disease — 
hematogenous icterus — is distinguished by the presence of biliary 
coloring matters, and absence of biliary acids in the urine while 
in hepatogenous icterus, both coloring matters and biliary acids 
are found in the urine. 

TESTS FOR BILIARY COLORING MATTERS IN URINE. 
Icteric urine may be recognized by its physical properties. 
The color is dark yellow, sometimes brown, and, when shaken, 
forms a yellow foam. 

GMELIN'S TEST. 

Into a test tube one-fourth full of nitric acid containing some 
nitrous acid — yellow nitric acid formed by exposure to light — let 
run down the side of the test tube some of the urine, so that it 
will remain on the acid. This is effected by holding the test tube 
containing the acid as near horizontal as possible, while the urine 
runs in from another test tube. The point of contact of urine 
and acid will undergo several changes in color, beginning with 
green and ending with yellow. The green color which appears 
in the reaction is characteristic of biliary coloring matters. A 
modification of this test is to filter, dry the filter paper carefully, 
and by means of a glass rod to place a drop of nitric acid on 
the paper. A colored ring will soon form, enclosing the place 
moistened by the acid. In case the coloring matters are in small 
quantity, introduce about 50 cc. of the urine into a glass-stoppered 
100 cc. cylinder, render acid with acetic acid, and shake some 
time with about 20 cc. chloroform. Draw off the chloroform 
solution by means of a syphon or pipette, and introduce into a 



60 



CHEMICAL ANALYSIS OF THE URINE. 



ioo cc. glass cylinder containing 50 cc. water, in which 5 or 10 
grms. sodium carbonate are dissolved. After shaking some time, 
draw off the aqueous solution and test by employing Gmelin's 
test. 

COLORING MATTERS OF THE BLOOD. 
Haemoglobin is the coloring matter of the red corpuscles of the 
blood. By heat it is decomposed into haematin and albumen. 
A solution of haemoglobin is dark violet in color. It absorbs 
oxygen when exposed to the air, forming oxyhaemoglobin, which 
imparts a deep red color to its solutions. Haemoglobin and its 
oxidation product have different optical properties. A solution of 
haemoglobin placed in a glass cell with parallel walls (a, Fig. 3), and 



Table i. 

D E b 



1 


\ 1 




2 






1 






4* 




1 



rays of light from a lamp passed through the solution, will reveal, 
in the spectroscope, a broad band, as shown by Spectrum 2, Table 
1, between D and E; but as haemoglobin absorbs oxygen, the spec- 
trum of the oxygen compound — oxyhaemoglobin — may likewise 
be seen, which consists of two bands or lines, one yellow and the 
other green, between D and E, Spectrum 1, Table 1. By reducing 
oxyhaemoglobin, when in solution, by means of a few drops of 
ammon. sulphide, and examining the solution with the spectro- 
scope, the broad band, Spectrum 2, will be seen, with the absence 
of the yellow and green bands, Spectrum 1. 

By the action of acid salts on a solution of either haemoglobin 
or oxyhaemoglobin, it changes to methaemoglobin, a solution of 
which, by examination with the spectroscope, yields a well-defined 



TESTS FOR COLORING MATTERS IN URINE. 



Gl 



red line between C and D, Spectrum 3, Table 1. When the 
coloring matter of the blood is in the urine, this is the usual form 
in which it is found; but as methaemoglobin in urine, undergoing 
fermentation, changes to haemoglobin, and as the latter changes 
to oxyhemoglobin by exposure to the air, all three bodies may 
be found in urine. By heating a neutral or acid solution of either 
haemoglobin, oxyhemoglobin or methaemoglobin, the albumen 
formed by the decomposition coagulates, and is colored from 



Fig. 




[From Neubauer.) 

haematin. Decomposition of these bodies likewise takes place 
by the action of glacial acetic acid and common salt, when warmed 
in the absence of water. As haematin is liberated by heat, it is 
not found in urine containing the coloring matters of the blood. 



TESTS FOR COLORING" MATTERS OF THE BLOOD IN URINE. 
When coloring matter of the blood is in the urine, it is either 
enclosed in red corpuscles, as in haematuria, or in solution in the 
urine, haemoglobinuria, produced by the action of certain toxi- 



62 CHEMICAL ANALYSIS OF THE URINE. 

cants, as the mineral acids, potassium chlorate, and the poison of 
scarlet, and other infectious fevers. The different parts of the 
spectroscope are adjusted by first removing the prism, P (Fig. 3), 
and placing the tube, A, that light from the lamp passing through 
the slit illuminate the margins of the tube, which is determined 
by looking through it toward the lamp. The telescopic part 
of the apparatus, B, is adjusted that objects at a distance are 
distinctly seen. The prism is now placed in position, and the 
reflecting tube, C, is adjusted that light be reflected through the 
telescope, and the scale become visible by the observer at d. By 
excluding light from the prism by a black cloth placed over it, 
and with the admission of a bright light into the reflecting tube, 
the apparatus is ready for use. A glass cell with parallel walls, 
a, is filled with urine for examination. If the urine is alkaline, 
render it acid with acetic acid. The urine, if turbid, is filtered, 
and the filtrate is examined, but if no spectrum indicating the 
presence of coloring matters of the blood is observed, the urine, 
before filtration, is examined. Highly-colored urine may be 
treated with finely-pulverized basic lead acetate until a solution 
of it ceases to produce a precipitate, and, after mixing well by 
shaking, render alkaline with ammon. hydrate, filter, and examine 
the filtrate with the spectroscope. The filtrate contains any oxy- 
hemoglobin and haemoglobin that may be in the urine, and the 
precipitate contains the methsemoglobin. If the result of the 
examination of the filtrate is unsatisfactory, wash the precipitate 
with water, transfer to a beaker by means of a fine stream of 
water from a wash bottle, and treat the precipitate suspended in 
water with a solution of sodium carbonate. The lead compound 
of methaemoglobin is decomposed by sodium carbonate with the 
formation of lead carbonate and methsemoglobin set free. Filter, 
and examine the filtrate with the spectroscope. In case the 
urine contains biliary coloring matters in sufficient quantity to 
interfere with the spectroscopic examination for the coloring mat- 
ters of the blood, the former are removed by rendering the urine 
slightly alkaline with ammon. hydrate, and adding a solution of 
calcium chloride until, after mixing well by stirring, a precipitate 
ceases to form, when the filtrate is examined with the spec- 
troscope. The spectra of the coloring matters of the blood are 
described above. 



BLOOD IN URINE. 63 

HELLER'S TEST. 

Not having a spectroscope at hand, the chemical tests will 
answer the purpose. Render the urine strongly alkaline with a 
solution of sodium hydrate and heat to the boiling point. The 
precipitate formed is composed of calcium and magnesium phos- 
phates, colored red by hsematin. This test alone is generally 
sufficient, but to examine further, filter, wash the precipitate with 
a small quantity of water and dry at ioo° C. Introduce the dried 
mass into a clean, dry test tube, add a small crystal of common 
salt and treat with some glacial acetic acid, heat to the boiling 
-temperature, filter a small quantity (i cc.) through a very small 
filter into a watch glass, evaporate on a water bath at 50 to 6o° 
C.,* and examine the residue with the microscope. Crystals of 
haematin (Fig. 4) are found if coloring matter of the Fig. 4 . 

blood is in the urine. In order to employ the _.^v. 

microscopic test, without regard to the red color ,\k^ -^ ^^ 
of the precipitate, with sodium hydrate, the color- -? <0\ fj 

ing matter is separated (Struve) by rendering the ' 

urine alkaline with a solution of sodium hydrate, when a solution 
of tannic acid is added until a precipitate ceases to form ; then ren- 
der acid with acetic acid, filter, wash with water, dry, and transfer 
to an agate mortar. The dried mass is now triturated with a small 
crystal of sodium chloride, when it is transferred to a dry test 
tube and boiled with some glacial acetic acid. Filter through a 
small filter into a watch glass and evaporate to dryness at a low 
temperature. The residue is examined for haematin with the 
microscope. 

jBLOOD IN URINE. 
If blood is in the urine besides haemoglobin or its derivatives, 
the urine is examined for red corpuscles, albumen and fibrin. 
Urine containing blood is usually red in color, yet the color 



* For this purpose the method of evaporating at a low temperature, first suggested 
by Streng, may be employed. Place a circular piece of glass over an evaporating 
dish containing water, the margin of the glass no L extending more than 5 mm. over 
the dish. The water in the dish is heated to the boiling temperature, and a watch 
glass or slide, in or on which is the solution to be evaporated, is placed on the glass 
plate. Evaporation will take place slowly. To evaporate at a still lower tempera- 
ture, introduce a piece of thin paper between the watch glass or slide and the glass 
plate. 



64 CHEMICAL ANALYSIS OF THE URINE. 

varies from red to dark brown, depending on the quantity of 
blood present and on the condition of the urine, whether fresh or 
undergoing fermentation. 

To determine the presence of red corpuscles of the blood, 
refer to Urinary Sediments, Chapter v, and for albumen, refer to 
page 46. Albumen is always found in small quantity in urine 
containing haemoglobin or its derivatives, but if red corpuscles of 
the blood are present, the quantity of albumen is greater coming 
from the liquor sanguinis as well as from decomposition of the 
coloring matters of the blood. If, however, the number of red 
corpuscles is not great, and the quantity of albumen is consider- 
able, the indication is that the excess of albumen originates from a 
diseased condition of the kidneys producing albuminuria. If albu- 
men, separated by heating some of the urine acidified with acetic 
acid to the boiling temperature, comes from the blood proper in the 
urine, it will rise to the surface in the form of highly-colored flakes. 
Fibrin usually appears as small coagulae suspended in the urine. 
To separate them from the urine, filter through fine muslin and 
wash with cold water. Treat a part of the mass supposed to be 
fibrin with a dilute solution of sodium hydrate; if insoluble, the 
indication is that it is fibrin ; albuminous bodies dissolve in 
sodium hydrate. Treat another portion with a weak solution of 
sodium carbonate (1 part Na 2 C0 3 dissolved in 100 parts water). 
Fibrin dissolves completely in this solution if warmed gently 
several hours on a water bath. This solution is then filtered 
and tested with Millon's reagent, page 44, when a deep red color 

is produced. 

LEUCIN. 

Leucin and tyrosin are found in the urine as a result of cer- 
tain pathological processes, particulaily in acute yellow atrophy 
of the liver and poisoning by phosphorus. The formula of 

leucin is — 

C 6 H 13 N0 2 . 

It is an oily-like solid crystallizing in spherical bodies, a, Fig. 5. 
It is partly soluble in water, slightly soluble in alcohol and insol- 
uble in ether. It is not found in urinary deposits. 

TEST FOR LEUCIN IN URINE. 

One litre of fresh urine is treated with an excess of basic lead 
acetate. Filter, and precipitate the lead from the filtrate with 



TYROSIN. 



Qo 



Fig. 5. 



sulphuretted hydrogen gas. The lead sulphide is separated from 
the solution by filtering, when the solution is evaporated to dry- 
ness on a water bath, and the residue 
treated with strong alcohol several 
times, and the alcohol washings are 
filtered each time into a beaker. 
Evaporate the alcohol solution at a 
low temperature, and examine the 
crystals which separate from solution 
during evaporation, when, if leucin 
is present, the form of crystals as 
shown by a, Fig. 5, will be recognized. 

Place some of the crystals on a piece of platinum foil and heat 
gradually (Scherer); if leucin is present, a spherical, oil-like glob- 
ule will form which does not adhere to the foil. 




TYROSIN. 



The formula of tyrosin is- 



In water it is slightly soluble, and in absolute alcohol and ether 
it is insoluble. In a solution of potassium, sodium or ammonium 
hydrate or carbonate it dissolves. It is also soluble in dilute 
acids. Tyrosin crystallizes in needle-like crystals, b, Fig. 5. 
With Millon's reagent (page 44) a hot water solution of tyrosin 
yields a dark-red color (R. Hoffmann). With an excess of con- 
centrated sulphuric acid gently warmed, a light- red color is pro- 
duced, which changes to a violet red by the addition of a solution 
of ferric chloride (Piria). By carefully evaporating a solution of 
tyrosin with nitric acid in a small porcelain dish, a deep yellow 
color is produced, which changes to dark red by the addition of 
sodium hydrate (Scherer). 

TEST FOR TYROSIN IN URINE. 

Tyrosin is sometimes found as a sediment, refer to page 77. 
Tyrosin is separated from the urine by the process employed in 
separating leucin, but being less soluble in strong alcohol than 
leucin, it remains undissolved by treating with absolute alcohol. 
The residue containing it is treated with weak alcohol heated to 
the boiling temperature, and filtered while hot. By evaporating 
5 



60 CHEMICAL ANALYSIS OF THE URINE. 

the filtrate, crystals of tyrosin separate, which are examined by 
the microscope. They are represented by b, Fig. 5. For further 
proof, employ Hoffmann's and Scherer's tests. 

FAT. 
In the course of a great many pathological processes fat 
appears in the urine in some cases in the form of oil drops on the 
surface of the urine, and again as an emulsion ; in either form the 
urine is turbid and does not become clear by heating. In health, 
fat sometimes appears in the urine, when there is an excess of it 
in the blood from the ingestion of fats. Resulting from degenera- 
tions of the kidneys, liver and other organs, as brought about by 
cancer, phosphorus poisoning, gangrene, Bright's disease, yellow 
atrophy of the liver, fractures of the bones and occasionally from 
diabetes mellitus, fat appears in the urine. Lipuria is therefore 
symptomatic of various diseases. In chyluria, fat with albumen, 
lecithin and cholesterin is in the urine in the emulsified form. 
The urine is white and is more or less homogeneous. This 
disease depends on the presence of microscopic organisms in the 
blood — the Filaria sanguinis. 

TESTS FOR FAT IN URINE. 

The presence of fat in the urine, when enclosed in albuminous 

casts and epithelial cells, is determined by microscopic examination. 

Oil drops and globules of fat are represented by Fig. 6. In 

chyluria the fat is in the emulsified condition, and its 

q ' presence is determined without difficulty by micro- 

n£V>v scopic examination. If fat is in the form of drops 

-. floating on the surface of the urine, its presence may 

^ be determined by causing the absorption of the drops 

by white paper, and by drying the paper the fat is 

found to be fixed, and the parts of the paper containing the fat 

are difficult to wet with water. To separate fat from the urine, 

shake about 400 cc. urine in a 500 cc. glass-stoppered cylinder 

with 50 cc. ether and decant the ether into a distilling flask. 

This process may be repeated several times ; when the ether is 

distilled on a water bath, the residue is treated with ether and 

the ether solution is introduced into a high beaker and evaporated. 

Fat in the residue is tested by its insolubility in water, by the 



O 




CHOLESTERIN AND LECITHIN. . 07 

paper test and by heating in a platinum dish, when a penetrating 

odor is produced. Instead of shaking the urine with ether, 

Dreschel's apparatus, Fig. 7, for the extraction of fats, may be 

employed for this purpose. Evaporate one litre of 

urine on a water bath to dryness, mix the residue 

with an insoluble substance, as calcium or barium 

sulphate, introduce the mixture into a folded filter, 

which is placed into the flask, b, while the flask, a, 

is filled about one-half full with ether, and the tube, 

a, is connected with a condenser. 

The ether is distilled by heating the flask, A, on a 
water bath, when the vapor will pass up the side 
tube, and when condensed in the condensing tube, 
the ether will percolate through the mixture in the 
filter, dissolving the fat. By continuing the process 
thirty or forty minutes, the fat will be dissolved. 
The ether solution is transferred to a high beaker and evapo- 
rated to dryness, when the presence of fat in the residue is deter- 
mined as above. 

CHOLESTERIN AND LECITHIN. 

Cholesterin, C24H44O, is a crystalline body, forming large tabular 
crystals. It is insoluble in water, soluble in ether and chloroform. 
When a chloroform solution of cholesterin is treated with an 
equal volume of strong sulphuric acid, it turns yellow ; with an 
excess of cholesterin the color is red or purple (Salkowsky). 
Cholesterin undergoes no change when a solution of it is heated 
with potassium, sodium, ammonium or barium hydrate. 

Lecithin, C42H 84 NP0 9 , is a solid easily decomposed by heat. It 
is insoluble in water, soluble in ether and chloroform. When a 
solution of lecithin is heated with barium hydrate or hydrates of 
the metals of the alkalies, it decomposes into neurin and glycerin- 
phosphoric acid. 

The method (Hoppe-Seyler and Eggel) employed for deter- 
mining the presence of cholesterin and lecithin in urine is based 
on their properties given' above. As these bodies are never found 
in large quantities in urine, they with fat are separated from four 
or five litres of urine. The urine, in portions of 500 or 1000 cc, 
is shaken up with ether after having been acidified, if not already 



68 CHEMICAL ANALYSIS OF THE URINE. 

acid in reaction. The ether solution having been drawn off by 
means of a syphon, the urine is rendered alkaline with a solution 
of sodium hydrate, and shaken with ether in the same way. The 
ether solutions of both acid and alkaline urine are distilled on a 
water bath, the residue treated with ether, free of water, and the 
solution is transferred to a 400 cc. flask. The ether having been 
distilled from the 400 cc. flask, the residue is boiled about two 
hours in 200 cc. of a saturated solution of barium hydrate, with 
a condenser connected with the flask, placed at about 35 degrees, 
so that the water condensed will return to the flask. At the end 
of the process, the fat will be decomposed into glycerin and fatty 
acids ; the latter forming an insoluble soap with barium. Lecithin 
will be decomposed into neurin and glycerin-phosphoric acid, 
the latter forming a salt with the barium present, while cholesterin 
remains unchanged. The residue, cholesterin and barium soap, 
is separated by filtering, and, having been washed with some 
water, it is transferred to a 50 cc. glass cylinder, treated with 
ether in small portions, and after shaking with each portion, the 
ether solution is decanted from the residue into a high beaker. 
The residue is finally treated with a mixture of absolute alcohol 
and ether, and the solution added to the ether solution in the 
beaker, when, by slow evaporation, cholesterin will crystallize 
out. Examine the crystals with the microscope; the crystals 
are tabular in form. Prepare a chloroform solution and test 
with strong sulphuric acid, as above. 

Neurin, one of the products of the decomposition of lecithin, 
is in solution in the filtrate from the residue of cholesterin and 
barium soap. 

To separate barium hydrate, which is also in the filtrate, pass 
carbonic acid gas through the solution, and separate the BaC0 3 
by filtering. 

Evaporate the filtrate on a water bath to dryness and treat the 
residue with absolute alcohol. The neurin will dissolve in alcohol, 
and the glycerin-phosphate of barium will remain undissolved. 
Filter through a dry filter paper and treat the filtrate with an 
alcohol solution of platinum chloride containing some hydro- 
chloric acid. A precipitate forms, composed of platinum chloride 
and neurin. Filter, and wash with a small quantity of alcohol, 
and dissolve on the filter with water. The aqueous solution is 



CHOLESTERIN AND LECITHIN. * 69 

concentrated by evaporation at a low temperature, better in vacuo, 
when the neurin compound will crystallize in six-sided crystals, 
known by microscopic examination. 

The presence of glycerin in the glycerin-phosphate of barium, 
from which the neurin was dissolved with absolute alcohol, is 
determined by drying some of it, and heating in a small porcelain 
dish, when the odor of acrolein will be perceptible. The odor of 
this body is characterized by its irritating properties. The test 
for glycerin is made with greater certainty (Hoppe-Seyler) by 
triturating some of the dried substance with acid potassium sul- 
phate, and heating the mixture in a dry test tube. 



CHAPTER V. 

Sediments in Urine — Sediments Peculiar to Acid Urine — The Acid Urates and Uric 
Acid — Hippuric Acid — Calcium Sulphate — Sediments Peculiar to Urine of Strong 
Alkaline Reaction — Calcium and Magnesium Phosphates and Magnesium Ammon. 
Phosphate — Ammonium Acid Urate — Calcium Oxalate — Calcium Carbonate — 
Sediments not Depending on Reaction of the Urine — Cystin — Tests for Cystin in 
Solution in Urine — Cystin as a Sediment — Tyrosin — Epithelial, Albuminous and 
Blood Casts — Blood Corpuscles — Pus — Spermatozoa, Bacteria, Sarcina, and Other 
Microorganisms. 

SEDIMENTS PECULIAR TO URINE OF STRONG ACID REACTION. 
THE ACID URATES AND URIC ACID. 

For the constitution and properties of uric acid and the urates 
refer to page 22. This sediment is usually red, yellow or "brick 
dust" in color. By microscopic examination, the acid urates are 
found to be amorphic or granular, seldom crystalline, except am- 
monium urate, which is sometimes present. A sediment com- 
posed of the urates disappears by heating the urine in which it is 
suspended and reappears as the urine cools, the acid urates being 
more soluble in hot than in cold water. Separate the sediment 
composed of urates from the urine by filtering, wash with a small 
quantity of cold water, transfer to a test tube, mix with some 
water and add a solution of sodium hydrate or carbonate; the 
acid urates will dissolve, owing to the formation of neutral urates, 
which are quite soluble. For this reason the acid urates may 
form as a sediment in urine of acid reaction^ but not in urine 
alkaline by the presence of potassium or sodium carbonate. 
Besides the solubility of the acid urates, by warming the urine 
containing the sediment, the granular form and the red color of 
the sediment, the urates may be still further tested by deter- 
mining if they are soluble in a solution of sodium hydrate with 
the microscope. For this purpose a small quantity of the sodium 
hydrate solution is brought on the margin or at one side of the 
glass cover, when the granular matter dissolves as the alkaline 
solution comes in contact with it. 

Uric acid, as a sediment in fresh urine, is not of such frequent 
occurrence as in urine having stood some time. By standing, the 
acid phosphates react on the neutral and acid urates, combining 

70 



SEDIMENTS PECULIAR TO URINE.- 71 

with some of their bases, and liberate more or less uric acid. For 
this reason the quantity of free uric acid in the sediment of 
urine having- stood several hours depends on the acidity of the 
urine and the relative amount of urates present. As this is gen- 
erally a slow process, the crystals of uric acid are well formed. 
The forms of the crystals of uric acid as generally found in uri- 
nary sediments, and the amorphic or granular character of the 
urates, are represented by Fig. 8. Uric acid, separated from com- 
bination by means of an acid, crystallizes in forms different than 
when spontaneously separated in the urine. The forms of 
crystals of uric acid separated by an acid are represented by 
Fig. 9. It is important, as regards the danger of the formation 
of concretions in the bladder, to ascertain if in urinary sediments 
of fresh urine there is uric acid, or if it forms by standing before 
the urine loses its acidity. To separate uric acid from the urates 

Fig. 8. Fig. 9. Fig. 10. 



in a sediment, heat the urine with sediment, filter while hot, and 
wash with hot water. The residue is tested with nitric acid and 
ammon. hydrate, page 23. A heavy deposit of urates and uric 
acid is no evidence of an increased quantity of uric acid in the 
urine, for, if there is present no excess of the acid phosphates, the 
bases may be in sufficient quantity to form neutral urates, and 
hence uric acid be in great excess without the formation of a 
sediment containing urates or uric acid ; therefore, to determine 
if uric acid is in excess in urine, quantitative estimations are 
made. 

HIPPURIC ACID. 

Hippuric acid is not of frequent occurrence in urinary deposits. 
The acid crystallizes in rhombic prisms and needle-like forms 
(Fig. 10), resembling the crystals of magnesium ammon. phos- 
phate. It differs, however, from the latter in appearing as a 
sediment in urine strongly acid in reaction, and by its insolubility 




VI CHEMICAL ANALYSIS OF THE URINE. 

in acetic acid, which is readily determined in the field of the 
microscope, with a drop of acetic acid placed at the margin of 
the glass cover. It sometimes occurs that crystals of hippuric 
acid, needle-like in form, are attached to crystals of uric acid 
when the latter are larger in size than are generally found in 
urinary deposits. Hippuric acid is readily distinguished from 
uric acid by its solubility in warm water and also in acetic ether. 

CALCIUM SULPHATE. 

The conditions for the formation of calcium sulphate as a 
sediment are concentration of the urine and strong acid reaction. 



Fig. ii. 



Fig. 12. 




Fig. 13. 




There are but a few cases reported of calcium sulphate having 
been found as a sediment in urine. Its crystals are needle-like 
and prismatic in form, as represented by Fig. n. 

SEDIMENTS PECULIAR TO URINE OF STRONG ALKALINE REACTION. 



CALCIUM AND MAGNESIUM PHOSPHATE AND MAGNESIUM AMMONIUM PHOS- 
PHATE. 

Although alkaline urine is generally turbid when fresh, it is 
sometimes nearly clear, when by standing or gently warming it 
becomes cloudy, a thin film forming on the surface before a 
deposit forms. The basic phosphates of calcium and magnesium, 
as found by the microscope in urinary deposits, are amorphic, 



SEDIMENTS PECULIAR TO ALKALINE URINE. 73 

and the granules or particles are sometimes so fine that they are 
seen with difficulty, unless glasses of high power be employed. 
The granular character of this sediment is illustrated by Fig. 12. 
Well-formed crystals of basic magnesium phosphate have been 
found in urinary deposits, but they are of rare occurrence. They 
are long, tabular in form, with pointed extremities. On the 
other hand, crystals of magnesium ammonium phosphates are 
frequently found. The crystals are usually well formed, large in 
size, rhombic in form, and resemble, somewhat, a coffin lid (Fig. 
13). As there is a comparatively small amount of ammonia salts 
in normal urine, unless it has undergone fermentation, the quan- 
tity of this salt is limited. The phosphates are quite soluble in 
acetic acid. They do not dissolve by heating the urine in which 
they are suspended or by the addition of potassium or sodium 
hydrate. By placing a drop of acetic acid or a solution of 
sodium hydrate at the margin of the glass cover, under which 
there is some of the sediment, the action of either of these 
reagents is determined with the microscope, or, to determine if 
the sediment is soluble in the urine when warmed, the slide may 
be carefully warmed, and the effect, if any, noted while the pre- 
paration is still warm. The mere fact that in fresh urine these 
phosphates form a sediment, is of no importance, for they sepa- 
rate from normal urine when alkaline and normal urine becomes 
alkaline by a vegetable diet. All of the phosphoric acid in alka- 
line urine is not combined with calcium and magnesium, but 
more or less remains in solution combined with potassium and 
sodium. It is important, however, to ascertain if the urine con- 
tinues alkaline even by varying the diet, and if so, microscopic 
examination of the urine is made to determine if micrococcus 
ureae are present, which is evidence of fermentation of the urine. 
If the alkaline reaction is due to ammonium carbonate, red lit- 
mus paper, when wet with the urine, will turn blue, but when 
dried the red color will be restored, or, by bringing the end of a 
glass rod, wet, with strong hydrochloric acid, near the surface of 
the urine, a cloud will form where the vapor of ammonium car- 
bonate comes in contact with the acid. 

In urine neutral, slightly alkaline, or acid, the neutral phos- 
phate of calcium — 

CaHP0 4 , 



74 CHEMICAL ANALYSIS OF THE URINE. 

sometimes appears as a sediment, the crystals of which are lance 
or wedge shape, often forming rosettes (Fig. 14). 

AMMONIUM ACID URATE. 
Ammonium acid urate appears as a sediment, the result of 
fermentation, when the ammonia formed decomposes the urates 
of potassium and sodium with the formation of ammonium acid 
urate. It is nearly insoluble in water. Its crystals are spherical in 
form, some having fine protuberances resembling the wild goose- 
berry (Fig. 15). In urine not containing an excess of ammonia, 
the NH 4 combines by preference with phosphoric acid and mag- 
nesium, forming MgNHJPQi, rather than with uric acid. It is 
only in urine containing ammonia from fermentation, or urine to 
which an ammonium compound has been added, that ammonium 

Fig. 14. f Fig. 15. 



--■/' 




Fig. 16. 



<g> ♦ 



u 



& E 



urate appears in considerable quantity as a sediment. Dilute 
hydrochloric or acetic acid decomposes ammonium urate with 
the separation of uric acid. Uric acid separated in this way is 
represented by Fig. 9, page 71. 

CALCIUM OXALATE. 
Calcium oxalate crystallizes from neutral or alkaline urine in 
octahedral crystals, and by the microscope the margins of the 
crystals, as they cross, are seen, which suggested the idea of the 
appearance of an envelope (Fig. 16). Occasionally the crystals 
are dumb-bell or spherical in form. The crystals of calcium 
oxalate are exceedingly small in size, and when not accompanied 
by other bodies may escape detection. If the sediment is small 
in quantity, filter the urine in which the sediment is suspended 



CALCIUM CARBONATE. 75 

through a small Swedish filter paper, wash with some water, 
transfer to a small beaker by means of a fine stream of water 
from a wash bottle and a hair pencil, and examine the sedi- 
ment with a microscope. With crystals of calcium oxalate, 
calcium, and magnesium phosphates (granular sediment), ammo- 
nium urate (spherical crystals), and magnesium ammonium phos- 
phate (rhombic crystals) are usually found. To determine the 
presence of calcium oxalate in the sediment of urine strongly 
alkaline in reaction, filter as above and dissolve the phosphates 
on the filter with dilute acetic acid, and, having washed with 
water, transfer the residue to a small beaker, as above, and 
examine for calcium oxalate with the microscope. As calcium 
oxalate differs from the phosphates in being insoluble in acetic 
acid, and from the urates (acid) and uric acid, in being insoluble 
in a solution of sodium hydrate, these reagents are employed 
to distinguish the oxalate from these sediments. The reactions 
may be made on the glass slide by placing a drop of the reagent 
at the margin of the glass cover, and observing what takes place 
as the reagent passes in the field of view. The quantity of oxalic 
acid in the urine is not increased by any disease, as far as has 
been determined, and regarding the existence of oxaluria path- 
ologists do not agree. In nearly all the researches made on this 
subject, the oxalic acid combined with calcium as a deposit in 
alkaline urine, as found by microscopic examination, has been 
taken into account, but no quantitative estimations were made. 
As the question turns on quantity rather than the presence of 
oxalic acid in the urine, investigations made so far are of but 
little value. 

CALCIUM CARBONATE. 
Calcium carbonate is not often found as a sediment. It 
generally forms by a continuous vegetable diet, when the urine 
becomes alkaline. From the urine it generally separates in 
spheroidal crystals, which occasionally become connected, so as to 
resemble dumb-bells. The forms of crystals, as generally found 
in urinary sediments, are represented by Fig. 17. Calcium 
carbonate dissolves in dilute acetic acid with the liberation of car- 
bonic acid gas. If the quantity of calcium carbonate in a sediment 
is small, this reaction is made in the field of the microscope by 
placing a drop of the acid at the margin of the glass cover, and when 



7b CHEMICAL ANALYSIS OF THE URINE. 

it comes in contact with the crystals, gas will evolve, leaving no 
doubt as to the constitution of the crystals. 

SEDIMENTS NOT DEPENDING ON REACTION OF THE URINE. 

CYSTIN. 

The formula of cystin is C 3 H 7 NS0 2 . It is seldom found in 
urine, and its presence is pathognomonic of no particular disease. 
It is found mostly in sediments and concretions. Cystin is 
insoluble in water, ether, alcohol and acetic acid ; soluble in 
hydrochloric acid and a solution of potassium, sodium or ammon. 
hydrate. When cystin is dissolved in a solution of sodium 
hydrate, and boiled with some lead hydrate, a black precipitate 
is formed — lead sulphide; the cystin is decomposed, and the 
sulphur in it combines with the lead. When some sodium nitro- 

Fig. 17. Fig. 18. 




prusside is added to a hydrate of sodium solution of cystin, the 
solution turns violet. Cystin crystallizes in six-sided prisms 
(Fig. 18). 

TESTS FOR CYSTIN DISSOLVED IN THE URINE. 

Treat 500 cc. of the urine (Loebisch) with an excess of acetic 
acid ; filter, wash with water and dissolve on the filter with water 
containing ammon. hydrate. Evaporate the filtrate in a beaker 
on a water bath, and cystin will separate from solution in crystals. 
Examine the crystals with the microscope and prepare a hydrate 
of sodium solution, and test with a small quantity of a solution of 
sodium nitroprusside, when the solution becomes purple in color, 
if cystin is present. 

CYSTIN AS A SEDIMENT. 

Filter the urine containing the sediment, wash with water, and 
dissolve on the filter with water containing ammon. hydrate. 



EPITHELIAL, ALBUMINOUS AND BLOOD CASTS. 7/ 

Render the ammonia solution distinctly acid with acetic acid and 
separate the cystin which precipitates by filtering, when some of 
the precipitate is dissolved in a solution of sodium hydrate and 
tested with a small quantity of sodium nitroprusside. If the solu- 
tion turns purple or violet in color, cystin is present. 

TYROSIN. 

Tyrosin, when in a sediment, is separated from other constituents 
of the deposit by the method employed above in separating 
cystin. Crystals of tyrosin precipitated by acetic acid are 
dissolved in dilute amnion, hydrate, to which some ammon. 
carbonate has been added, filtered, if necessary, and evaporated 
at a low temperature, when tyrosin will separate in the form of 
crystals, b, Fig. 5, page 65. Examine the crystals with the 
microscope, and test according to R. Hoffmann and Scherer, 
page 65. 

EPITHELIAL, ALBUMINOUS AND BLOOD CASTS. 

The sediment of urine containing albumen generally contains 
small cylindrical fragments, which are formed in the uriniferous 
tubes, and hence are known as casts or tube casts. Besides 
blood casts, composed of blood corpuscles and fibrin, the result 
of hemorrhage, there are three kinds of casts, epithelial, hyaline 
and waxy. Epithelial casts are composed of the epithelial cells 
of the canals or tubes in which they are formed. The epithelial 
cells of these tubes are round-like bodies, represented by Fig. 23, 
page 79. 

As the casts are made up of these cells, they are recognized 
by their dentated or irregular surfaces. In some of the casts the 
epithelial cells appear in rows, each having a well-defined outline. 
In other casts the rows are difficult to make out, and the outline 
or contour of each cell is indistinct; those of the latter class are 
known as metamorphosed epithelial casts. Both varieties are 
represented by a and b, Fig. 19. Epithelial casts refract light to 
a greater extent than the urine, hence with the microscope they 
are not difficult to find, as they appear bright with well-defined 
outlines. Epithelial casts are firmer in consistence, and resist the 
action of chemical reagents to a greater extent, than hyaline casts. 

Hyaline casts (Fig. 20) are smaller and have smoother surfaces 



78 



CHEMICAL ANALYSIS OF THE URINE. 



than epithelial casts. They are not colored and refract light less 
than the urine, and are, therefore, more difficult to find with the 
microscope. Those that are somewhat long are usually bent, 
while the short ones are straight. Hyaline casts are somewhat 
granular, the granules being metamorphosed constituents of the 
uriniferous tubes. Waxy casts are the largest in size. They 
refract light to a much greater extent than the urine, so that in 
the field of the microscope they appear bright. They are repre- 
sented by Fig. 21. They are slightly yellow in color. As casts 
readily disintegrate, especially in alkaline urine, the sediment is 



Fig. 19. 



Fig. 20. 



Fig. 21 




P 






i :.-%\? : :--: 




examined, if possible, as soon as it settles. In case of failure to 
find any casts in several preparations, filter some of the urine 
with sediment suspended, wash with a small quantity of water 
and transfer the sediment to a conical wine glass, and add a 
solution of either eosin or gentianin in quantity sufficient to 
impart a distinct color to the fluid, and after standing until the 
sediment subsides, it is examined with the microscope. The casts 
will be stained so as to become visible. Without having filtered, 
washed, or stained the sediment, and there is doubt as to the 
presence of hyaline casts, the process of staining may be carried 
on with the sediment on the slide (Rouvida). For this purpose, 



EPITHELIAL CELLS AND MICROORGANISMS. 



79 



water, in portions of one or two drops, is placed at the margin 
of the glass cover, and the fluid is absorbed from the opposite 
margin of the cover by means of slips of filter paper. Having 
thus washed the sediment, a small quantity of solution of gen- 
tianin is placed at the margin of the cover, and is drawn under 
the glass by a piece of filter paper, as in washing, when, in a short 
time, hyaline casts will become visible. 

BLOOD CASTS. 
Blood casts are formed of blood in the uriniferous tubes by the 
coagulation of its fibrin. They are, therefore, composed of red 
corpuscles and fibrin. They are represented by Fig. 22. As they 
appear in urine with blood, they are difficult to find, especially 
when the amount of blood is considerable. They are about the 



Fig. 22. 



Fig. 23. 



Fig. 24. 



Fig. 25. 




size of epithelial casts, but are dark, from the blood corpuscles 
they contain, 



EPITHELIAL CELLS, BLOOD CORPUSCLES, PUS, SPERMATOZOA, 
BACTERIA, AND OTHER ORGANISMS. 

Normal urine contains a few epithelial cells from the urethra, 
vagina of the female, ureters and pelvis of the kidneys. 

By diseases of the mucous membrane the number of epithelial 
cells in the urine is increased, and in chronic inflammatory or 
degenerative processes, as in Bright's disease, amyloid and fatty 
degenerations, epithelial ' cells from the parts diseased are often 
metamorphosed, that is, contracted, and often contain particles 
of fat. Epithelial cells from the uriniferous tubes and urethra of 
the male differ from those of the pelvis of the kidney and bladder, 



80 CHEMICAL ANALYSIS OF THE URINE. 

so that they may be recognized by the microscope. The epi- 
thelial cells of the uriniferous tubes are somewhat round and 
small (Fig. 23), while those of the pelvis of the kidney and bladder 
are flat and thin (Fig. 24). The epithelial cells of the neck of 
the bladder are irregular in shape (Fig. 25). 

BLOOD CORPUSCLES. 
Clots of blood may form in the urine after it has been passed, 
or they may form in the bladder and obstruct the passage of 
urine. Urine containing blood is generally cloudy and red or 
dark red in color, unless the quantity of blood present is small, 
when the urine is so changed in color that the presence of blood 
is suspected. In cases of hemorrhage of any part of the urinary 
passages not complicated by nephritis or cystitis, except at point 
of hemorrhage, the urine may be nearly normal, except that it 
contains blood. Generally, however, hemorrhages accompany 
acute inflammation of the kidneys, or they are produced by 
chronic ulcerative processes in the walls of the bladder, when the 
sediment contains pus and epithelial cells. The presence of 
blood casts in the sediment, a considerable quantity of albumen 
in the urine, and the absence of coagulae of blood, not distin- 
guishable without the aid of a lens or microscope, is evidence 
that the hemorrhage occurred in the kidneys. In urine con- 
taining blood or pus, fermentation begins much sooner than in 
normal urine, and as tube casts undergo changes in the presence 
of bacteria, by which they cannot be recognized, urine in which 
the presence of blood is suspected should be examined without 
delay. Blood corpuscles in the urine are often modified in form ; 
sometimes they are nearly round, sometimes dentated, and again 
shrunken and apparently atrophied. Blood corpuscles, as they 
generally appear, are represented by Fig. 26. If in case of doubt 
of the presence of blood in the urine by microscopic examination, 
employ the spectroscopic test for coloring matters of the blood, 
page 61, or, not having a spectroscope at hand, employ Struve's 
test for haematin, page 63. For the tests for fibrin, refer to 
page 64. 

PUS. 

Pus cells, as they appear in the field of the microscope, are 
round-like bodies about twice the size of the red corpuscles of 



pus. * 81 

the blood. They are represented by a, Fig. 27. The contents of 
pus cells are granular. They contain one or two nuclei, but on 
account of the granular matter they contain, the nuclei are not 
seen. By microscopic examination or by means of chemical 
reagents, mucus corpuscles cannot be distinguished from pus 
cells. In normal urine there are mucus corpuscles, but the 
number is limited. By irritation or inflammation of the mucous 
membrane of the urinary passages the number of mucus cor- 
puscles in the urine is greatly increased. But at what period or 
stage of the disease pus cells appear in the urine is in some cases 
difficult to determine. If the inflammation is not confined to a 
small surface, but involves the mucous membrane of the bladder, 
the urine is turbid, often containing shreds of mucus, and when 
filtered the filtrate yields a light precipitate, mucine, by the addi- 
tion of acetic acid. Besides pus, as found by microscopic exami- 
nation of the sediment, the urine contains a small quantity of 
albumen, to determine the presence of which the urine is filtered, 

Fig. 26. Fig. 27. Fig. 28. 




£>* 



mw 



and the filtrate rendered strongly acid with acetic acid, when it 
is filtered and the filtrate tested for albumen — refer to tests for 
Albuminous Bodies, page 46. If the quantity of albumen is 
considerable a part comes from the kidneys, and a careful exami- 
nation of the sediment is made for tube casts. If pus comes 
from cystitis and the urine is alkaline from fermentation having 
taken place, crystals of magnesium ammon. phosphate (a, Fig. 29) 
and ammon. urate (b, Fig. 29) will be found in the sediment. In 
this condition of the urine the presence of pus cannot be deter- 
mined with certainty by microscopic examination of the sediment, 
unless the urine has been alkaline but a short time. Alkalies 
change pus to a gelatinous mass, the cells dissolve leaving the 
nuclei. The contour of the cells is seen if the action of the 
alkali, ammon. carbonate in urine, undergoing fermentation, has 
not continued long. A similar change in pus cells takes place 



82 CHEMICAL ANALYSIS OF THE URINE. 

by the action of dilute acetic acid (b y Fig. 27). The change in the 
cells is seen by placing a drop of the dilute acid at the margin of 
the glass cover, and by means of a piece of porous paper causing 
absorption of fluid from the opposite margin of the cover, when 
the acid will pass under the cover and come in contact with the 
cells. The disappearance of the walls of the cells with the gran- 
ular matter is seen. The nuclei appear as dark, irregular frag- 
ments. The presence of pus in the sediment of urine, of acid 
reaction, is determined by stirring a piece of sodium hydrate with 
the sediment, when the pus becomes gummy and tenacious 
(Donne). In inflammation of the urethra of the male — gonorrhoea 
— the sediment of the urine contains pus in quantity, depending 
on the stage of the disease and frequency of passing the urine ; 
but for the examination of urinary sediments containing patho- 
logical products of the urethra, small quantities of the urine first 
passed are collected in a conical wine glass, the sediment of which 
is examined. 

Pus enters the urinary passages from abscesses in the pelvis, 
or parenchyma of the kidney resulting from obstruction, as stone, 
or from cancerous or tubercular formations. In these cases many 
pathological products may be found in the urine. 

SPERMATOZOA. 

Spermatozoa in the urine are found only by microscopic ex- 
amination. To examine urine for spermatozoa, that passed first 
in the morning should be taken, and after standing several hours 
the sediment is examined by the microscope. The movements 
of spermatozoa soon cease in the urine, and especially if the urine 
is strongly acid in reaction. The general appearance of sper- 
matozoa is shown by Fig. 28. They have a head and tail but 
no neck. 

BACTERIA. 

Normal urine when passed from the bladder and collected over 
mercury, having been boiled, will undergo no change for many 
months, providing the process is so conducted that no air comes 
in contact with the urine (Pasteur, Cazeneuve and Liron). By 
exposure to the air urine undergoes fermentation (page 20). 

In urine having undergone fermentation in the bladder, as is 
frequently the case in inflammation of that organ, the micrococ- 
cus ureae are found as in fermentation of the urine by exposure 



SARCINA AND MICROORGANISMS. 83 

to the air. It is doubtful if the germs of bacteria are in normal 
urine not having come in contact with the air ; but that they are 
often introduced into the bladder by the use of unclean catheters 
admits of no doubt, and in cases of chronic cystitis in which 
no catheter has been used, and yet fermentation of the urine 
takes place in the bladder, the germs may have found their way 
into the bladder by relaxation of the constrictor muscle of the 
urethra resulting from a diseased condition. 

The micrococcus ureae appear as thread-like bodies, some long, 
others short, and during life they are in continued movement. 
In alkaline urine they are generally found in the sediment with 
crystals of magnesium ammonium phosphate, ammonium urate 
and granular calcium and magnesium phosphates, as in Fig. 29. 

Fig. 29. Fig. 30. 






-%> 




. 



By a high magnifying power the thread-like fragments are found 
to be made up of minute cells. By infectious diseases, as scarlet 
fever, diphtheria, erysipelas, etc., the urine is found to contain 
microorganisms. The kidneys, in the process of separating 
these organisms from the blood, often become inflamed, giving 
rise to albuminuria as an accompaniment of the disease. On the 
other hand, it is claimed that the albuminuria which is developed 
in these cases is due to diminished blood pressure in the kidneys, 
the result of high fever. 

SARCINA AND' OTHER MICROORGANISMS. 
It is necessary that nutritive fluids be differently constituted 
for the culture of different microorganisms, so that the germs of 
those organisms for which the conditions of life are favorable 



84 CHEMICAL ANALYSIS OF THE URINE. 

will germinate. It is probable that various forms of fermentation 
would take place in the urine while in the bladder in the course 
of infectious diseases, if the urine were so constituted that the dif- 
ferent organisms excreted by the kidneys could be cultivated in 
it. Besides the micrococcus ureae the sarcina are cultivated in the 
urine, yet the latter are rarely found in the urine. They differ 
from the sarcina sometimes found in the stomach in being- about 
half the size. Urine containing sarcina is usually cloudy, and 
often contains a heavy sediment composed mostly of the organ- 
isms. Sarcina do not produce pathological changes in the 
mucous membrane of the urinary passages, neither is their pres- 
ence in the urine associated with any general disease. In the 
sediment they are easily found by microscopic examination, as 
they differ from other organisms by being composed of cells of 
4, 8, 1 6, 64 and sometimes 512 (Fig. 30). In the sediment of 
diabetic urine, cells of the ordinary yeast ferment are sometimes 
present. They are round-like bodies about the size of pus cells, 
but they have no nucleus and are usually in clusters. From 
pus cells they are distinguished by no nuclei appearing by the 
action of dilute acetic acid. 

Of the moulds, the penicilium glaucum is found in urine more 
frequently than any other. When in urine, it usually appears on 
the surface, at first as a colorless film, and in time it becomes 
green or blue and more dense. By microscopic examination it 
is found to be composed of a meshwork of fine fibres — myce- 
lium — as shown by Fig. 31, and with the fibres round cells, the 
spores, are generally found. 



CHAPTER VI. 

Scheme for the Qualitative Analysis of Healthy and Diseased Urine — Sediments — 
Microscopic Examination of Sediments — Staining — Scheme for the Qualitative 
Analysis of Sediments. 

QUALITATIVE ANALYSIS OF HEALTHY AND DISEASED URINE. 

In examining the urine in the order here given, frequent 
reference is made to the more elaborate consideration of the 
properties of each body, as well as to the methods employed in 
separating it from the urine, and identifying it in the preceding 
pages. This scheme embraces only those bodies in the urine a 
knowledge of which is of the greatest importance, and it is 
intended to serve the purpose of an index or guide in determining 
the quality of the urine, whether healthy or diseased. Following 
the qualitative examination of the urine will be found the scheme 
for analysis of urinary sediments, followed by methods employed 
in determining the constitution of concretions or stones. 

Determine if the urine, when fresh and before it cools, if 
possible, is clear or turbid, and if clear, if a sediment 

Properties f° rms as it cools. If fresh urine is turbid, determine 
if the sediment is amorphic or crystalline, and if 
crystals are present, their character — that is, if difficult to see 
without the aid of a glass, or if they are coarse (gravel). 

Urine, highly colored, may be normal, but if so, it is con- 
centrated, and for a given volume contains more 

Color. ... 

urobilin than urine generally does. Urine of high 
specific gravity, from a person having fever, is highly colored; 
but diabetic urine, also of high specific gravity, is generally nearly 
colorless. If urine is highly colored, deep red or brown, 
examination for blood is made ; if the color is dark yellow or 
green, and by shaking the urine a yellow green foam forms, the 
urine is examined for coloring matters of the bile. 

If the odor of the urine is well pronounced, determine if due 

to fermentation by microscopic examination for the 

micrococcus ureae. The presence of an excess of 

ammon. carb. from fermentation gives to urine a strong urinary 

85 



86 CHEMICAL ANALYSIS OF THE URINE. 

odor. If the odor of urine is peculiar, it may be due to the 

ingestion of asparagus, turpentine, cubebs, etc. Determine the 

specific gravity of the urine by means of a urinometer. 

Gravky! The following conditions for making this test accurately 

are taken into account. The markings of the scale 

should not be too close ; the vessel holding the urine should be 

wide enough to permit the instrument to float without touching 

its sides ; the temperature of the urine should be brought to that 

for which the instrument is graduated, and the instrument should 

be dried with a piece of cloth before using. 

If blue litmus paper changes instantly when brought in contact 

with the urine, it is strongly acid in reaction, and if 

Properties. re d litmus paper turns blue at once, or yellow turmeric 

paper dark red, the urine is strongly alkaline in 

reaction. If the urine produces no change in the color of either 

blue or red litmus paper, it is neutral in reaction. Occasionally 

urine changes both blue and red litmus paper purple, owing to 

there being in solution a mixture of the neutral phosphates, 

which are alkaline in reaction, and the acid phosphates, which 

are acid in reaction. (Amphotic Urine of Bamberger.) 

If the urine is alkaline, examine for the micrococcus ureae ; 
if absent, the alkaline reaction is due to the presence of potassium 
or sodium carbonate. 

Fat in the urine can scarcely be regarded as a sediment ; if in 

the form of oil drops, the fat appears on the surface of 

the urine, and may possibly escape detection when the 

sediment is examined with the microscope. If, on the other 

hand, the fat is in the emulsified form, its presence would be 

discovered in examining the sediment with the microscope. 

In all examinations of the urine the surface should be carefully 
inspected to detect the presence of any floating particles of fat, or, 
if the urine by standing continues cloudy, a drop should be 
examined with the microscope to determine if fat globules or 
granules are present. (Refer to Fat in Urine, page 66.) 

For ordinary clinical purposes the tests for serum albumen and 

globuline in urine are sufficient. It is only when 

" B q^° us special investigations in pathological conditions, which 

may be characterized by the appearance of more or 

less globuline, hemialbumose and peptone, that the presence of 



ANALYSIS OF HEALTHY AND DISEASED URINE. 87 

these bodies in urine is determined. Before testing for albumen, 
urine not clear should be filtered. Test for albumen with 
nitric acid, also with the sodium chloride and acetic acid test, 
page 46. 

There are many other tests for albumen in urine, but these 
answer every purpose. For the method for testing for hemial- 
bumose refer to page 47, for peptone page 47, and for globuline 
and serum albumen page 47. 

Urine containing an abnormal quantity of mucine is usually 
turbid, and with an excess of acetic acid the turbidity 

Mucine. 

is increased. To test for mucine, filter the urine 
through a double filter ; dilute the filtrate with an equal volume 
of water and render strongly acid with acetic acid. A light 
precipitate forms if mucine is present. 

Before testing for sugar, filter the urine, if necessary. Unless 

the urine contains a small quantity of sugar, there will 
urine b e no difficulty in determining its presence by means 

of Salkowski's modification of Trommer's test (page 50) 
or with Fehling's solution (page 50); but when urine contains 
reducible substances in quantity sufficient to produce a yellow 
flocculent precipitate of cuprous hydrate, and doubt arises as to 
the presence of sugar, employ the fermentation test (page 52). 
With some experience in purifying salts by recrystallizing with 
but little loss, and determining the melting point of a body, the 
test for sugar in the urine in small quantities with phenylhydrazin 
(page 53) is practicable. Not having the fermentation tubes or 
phenylhydrazin at hand, coloring matter, etc., may be removed 
from the urine with either lead acetate or corrosive sublimate 
(page 51) and the filtrate tested for sugar with Salkowski's modi- 
fication of Trommer's test and Fehling's solution. If the results 
should be unsatisfactory, separate the sugar from the urine by 
means of potassium hydrate or Bruecke's method (page 54) and 
test the aqueous solution for sugar. 

To test for. inosit, refer to page 55. Testing for this body 

may be omitted, except in cases of diabetes and 

Inosit. J ' ^ 

Bright s disease. 
For determination of the presence of biliary acids in urine 
Biliary refer to page 58, and for biliary coloring matters 
Constituents. em ploy Gmelin's test, page 59. 



88 CHEMICAL ANALYSIS OF THE URINE. 

To test for leucin and tyrosin refer to page 65, but, unless 
Leudnand tnere are symptoms indicating disease of the liver, 
Tyrosin. fa Q examination for the presence of these bodies may 
be omitted. 

In cases of chronic diseases of the intestinal canal, 

Indican. 

test for indican (page 27). 

Concentrate the urine after having filtered, if necessary, by 

evaporating on a water bath, render strongly acid 

with dilute hydrochloric acid, and after standing 

twenty-four hours uric acid will separate from the neutral urates. 

Filter, wash and test with nitric acid and ammon. hydr. (page 23). 

To determine the presence of chlorine combined with bases in 

the urine, render distinctly acid with nitric acid ; add 

Chlorine. ' J ' 

a solution of silver nitrate. A white precipitate forms 
if chlorine is present. 

A part of the phosphoric acid combined with bases will pre- 
cipitate by rendering the urine strongly alkaline with 

Phosphoric . . . 

Acid of the ammon. hydrate, lhe precipitate formed is com- 

Phosphates. J . . , 

posed of the basic phosphates of calcium and mag- 
nesium. To precipitate all of the phosphoric acid combined 
with bases, add ammon. hydrate to the urine until strongly alka- 
line, when an excess of a solution of calcium chloride is added. 
To test for phosphoric acid combined in the form 
of Glycerin- 1 of glycerin-phosphoric acid, employ Sotnischewsky's 
method (page 34). 

Sulphuric acid combined with bases is precipitated 

Sulphuric Acid . . . . . . 

combined by rendering; the urine distinctly acid with acetic acid 

with Bases. J & . ..... 

and adding a solution of barium chloride. 
Sui huric 4cid T° test f° r sulphuric acid in ester compounds, refer 
<&££*. to page 36. 

For ammonia in combination, test by employing 

Ammonia. 

alcohol and a solution of platinum chloride (page 39). 

URINARY SEDIMENTS. 
It is evident that if urine contains a sediment, it is not soluble 
in the amount of urine in which it has formed. Some sediments 
form when the urine changes in reaction, and others, again, are 
produced by fermentation. In some cases the sediment is com- 
posed of organized elements of the tissues, or they are pro- 



URINARY SEDIMENTS. Q\) 

duced by pathological processes. It is therefore apparent that in 
the study of urinary sediments a number of data are taken into 
consideration. In collecting urine for examination, vessels abso- 
lutely clean are employed, otherwise fermentation of the urine 
may take place in a short time, or fat and other bodies be found 
in the urine which came from the vessel. The sediment will 
separate from urine in a bottle or glass cylinder so that it may 
be examined, but if the quantity is small, a conical wine glass is 
preferred. Before examining the sediment the reaction of the 
urine is determined, and also if the urine was turbid or clear 
when passed. 

The principal part of the work of examining urinary sedi- 
ments is microscopic in character, and the conditions for secur- 
ing good results rest to a great extent on keeping clean the 
microscopic slides, glass covers, objectives, oculars — briefly stated, 
skill in working with the microscope, which, however, is not dif- 
ficult to acquire. 

The sediment having subsided, portions of it are drawn off 
by means of a glass tube about 25 cm. long, with internal diam- 
eter of four or five mm., and one end drawn out in the flame of 
a Bunsen's burner or spirit lamp, so that its orifice is about one 
millimetre in diameter. The upper end of the tube is fused in the 
flame of a blowpipe that its margins may be removed. By moist- 
ening the finger somewhat, and placing it firmly over the fused 
end of the tube, and sinking the drawn-out extremity into the 
sediment and partly removing the finger, air will pass out and 
some of the sediment with urine will enter the tube. The upper 
end of the tube is again closed and the sediment will remain in 
the tube, when a drop of the urine containing some of the sedi- 
ment may be placed on a slide for microscopic examination, or a 
larger quantity introduced into a test tube by removing the 
finger. Some sediments subside sooner than others, and con- 
sequently form the lower strata of the deposit, so that it is well 
to draw off portions of the sediment for examination at different 
distances from the bottom of the glass. About one drop of the 
mixture of sediment and urine is sufficient to fill the space 
between the slide and glass cover when the latter is placed on 
the fluid. The quantity should not be sufficient to float the glass 
cover or pass over its margins. Some chemical reactions may 



90 CHEMICAL ANALYSIS OF THE URINE. 

be made on the slide with the glass cover in place, more particu- 
larly those by which solution of solids takes place ; but when 
crystals which form by chemical reactions are to be examined, 
the reaction, as a rule, is brought about before the glass cover is 
placed on the preparation. Some of the urine with sediment is 
transferred to a test tube, a few drops of the reagent added and 
mixed with urine by carefully agitating the test tube, and the 
solution having remained in quietude one or two hours, a drop 
of the fluid with crystals is placed on the slide by means of the 
drawn-out tube. Crystals formed in this way are generally more 
fully developed than when formed on a slide, as the solution may 
become concentrated by evaporation before the crystals have 
formed, and with the glass cover in place the space may be 
insufficient. For the formation of fully-developed crystals, 
space for the free movement of the molecules (hence solutions 
should be as dilute as the case will permit) and absolute 
quietude are conditions which cannot be ignored. 

To ascertain if a sediment under examination dissolves when 
it comes in contact with a reagent, place a drop of the latter at 
the margin of the glass cover and bring a narrow strip of porous 
paper in contact with the fluid at the opposite margin of the 
cover ; the reagent will pass under the glass cover and come in 
contact with the sediment. Care is taken that the current estab- 
lished is not sufficient to carry the bodies out of the field of 
view. To find hyaline casts in urinary sediments, it may be found 
necessary to employ a staining fluid by which they become visible. 
For this purpose, after the sediment has subsided, one or two cc. 
of the fluid containing some of the sediment is drawn off with a 
pipette and introduced into a small clean test tube, and a solution 
of eosin added in quantity to impart a distinct color to the fluid. 
Mix well by gently agitating the test tube, and in fifteen min- 
utes transfer a drop of the fluid containing some of the sediment 
to a slide by means of the drawn-out tube. A solution of either 
picrocarmine or gentianin may be employed as well as eosin. 
These staining fluids may be purchased of any dealer in micro- 
scopic apparatus. The plan adopted here for examining sedi- 
ments is to test for the urates, etc., in the sediment of urine 
decidedly acid in reaction, and for the phosphates of calcium and 
magnesium, etc., in urine strongly alkaline in reaction (refer 



URINARY SEDIMENTS. 91 

above to reactions of the urine), and if in this course other bodies 
are observed, reference is made to methods employed in finding 
constituents of sediments not depending on the reaction of the 
urine for their formation. As the greater number of urinary sedi- 
ments do not depend on the reaction of the urine and cannot be 
separated into groups by reagents, any system for the analysis of 
them is necessarily defective. 

(A ) THE URINE IS STRONGLY ACID IN REACTION. 

1. Heat some of the sediment with urine in a test tube — it clears 

up and reappears by cooling. Presence of the urates and 
possibly hippuric acid. The sediment appears to remain 
unchanged. Boil gently a short time, filter while hot and 
place filtrate one side for several hours — it becomes cloudy 
and by warming clears up. Presence of the urates. The 
sediment is, therefore, partly composed of urates. 

2. By microscopic examination there is granular matter (Fig. 8, 

page 71), and when a drop of solution of sodium hydr. 
comes in contact with the granules, by placing it at margin 
of glass cover they dissolve. Presence of urates. 

3. The crystals are large and tetrahedral in form (Fig. 10, page 

71), soluble in ammon. hydrate. Presence of hippuric 
acid. 

4. The crystals are well formed, four or six sided (Fig. 8, page 71). 

They remain unchanged with dilute hydrochloric or acetic 
acid, but dissolve in a solution of sodium hydrate. Presence 
of uric acid. Uric acid may be further tested by filtering 
the urine containing the sediment, washing with hot water, 
and testing the residue for uric acid with nitric acid and 
ammon. hydr. (page 23). 

5. In the sediment there are crystals which are prismatic and 

needle-like in form, many radiating from a common centre 

(Fig. 11, page y 2). They are insoluble in hydrochloric acid. 

Presence of calcium sulphate. 

If, in the examination of the sediment of urine, strongly acid 

in reaction, by the methods here given, other bodies are observed 

by microscopic examination, to which no reference was made, 

examine them according to D and E ; and if there is reason to 

believe that the urine is not sufficiently acid in reaction to pre- 



92 CHEMICAL ANALYSIS OF THE URINE. 

vent the separation of the phosphates of metals of the alkaline 
earths, examine the sediment according to B, C and D. 

(B) THE URINE IS STRONGLY ALKALINE IN REACTION. 

1. By microscopic examination the sediment is granular or 

amorphic (Fig. 12, page 72). With acetic acid it dissolves. 
Presence of calcium and magnesium phosphates. 

2. The crystals are large, well formed, prismatic, some having 

no corners, resembling in appearance a coffin lid (Fig. 13, 
page 72). The crystals are soluble in acetic acid. Pres- 
ence of magnesium, amnion, phosphate. 

3. The crystals are spherical, sometimes, though seldom dumb- 

bell in form (Fig. 17, page j6). The crystals dissolve in 
dilute hydrochloric or acetic acid, and gas bubbles are 
formed under the glass cover. Presence of calcium car- 
bonate. 

(C) THE URINE IS NEUTRAL, SLIGHTLY ACID OR ALKALINE IN REACTION. 

I. By microscopic examination there are crystals wedge-shaped, 
having centres from which they diverge (rosettes), Fig. 14, 
page 74. The crystals are soluble in dilute acetic acid and 
insoluble in a solution of sodium hydrate. Presence of cal- 
cium neutral phosphate. With this sediment there may be 
bodies described under A, B and D. 

{D ) CRYSTALLINE BODIES ARE IN THE SEDIMENT NOT DEPENDING ON 
REACTION OF THE URINE. 

1. By microscopic examination the crystals are spherical, some 

having thorn-like outgrowths (as wild gooseberries), Fig. 
15, page 74. When treated with a drop of acetic acid they 
disappear, but crystals of uric acid form after the lapse of 
some time (Fig. 9, page 71). Presence of ammon. urate. 

2. The crystals are small,octahedral, with lines crossing (envelope 

form), Fig. 16, page 74. Occasionally they are in the form 
of dumb-bells. In dilute hydrochloric acid they dissolve, 
but are insoluble in acetic acid. Presence of calcium oxa- 
late. 

3. The crystals are six-sided and tabular (Fig. 18, page 76). The 

crystals undergo no change with acetic acid but dissolve in 
ammon. hydrate. Presence of cystin. 



URINARY SEDIMENTS. 93 

4. The crystals are needle-like and radiate from centres (rosettes), 
b, Fig. 5, page 65. The crystals are soluble in ammon. 
hydrate. Presence of tyrosin. Test the urine for leucin 
and tyrosin, page 65. 

(E) ORGANIZED BODIES— PATHOLOGICAL PRODUCTS— ARE IN THE SEDIMENT. 

1. By microscopic examination they are cylindrical bodies, hav- 

ing irregular but well-defined margins (a, Fig. 19, page 78). 
The cell formation of some is seen as shown by a, Fig. 19, 
while in others the surfaces are more homogeneous or agglu- 
tinated as seen by b, Fig. 19. Presence of epithelial casts. 
With dilute acetic acid they undergo no change. Those 
represented by b are metamorphosed. 

2. They are cylindrical in form, smaller than those of E I, sur- 

faces smooth ; contents homogeneous or granulated, with 
outlines so indistinct that they are seen with difficulty (Fig. 
20, page 78). Presence of hyaline casts. They dissolve 
slowly in acetic acid, and with iodine dissolved in a solution 
of potassium iodide they turn yellow. For the employ- 
ment of staining fluids in microscopic preparations, refer to 
page 90. 

3. They are cylindrical in form, larger than those cf E 1 or 2 ; 

contents amorphic gray or somewhat yellow in color, and 
more easily seen than those of E 2. Some are even in out- 
line, while others are tortuous (Fig. 21, page j8). Presence 
of waxy casts. 

4. They are cylindrical in form, dark in color, formed of round- 

like bodies (Fig. 22, page 79). Presence of blood casts. 
Examine the urine for coloring matters of the blood with 
the spectroscope (page 60) or by Struve's test (page 63), and 
for blood corpuscles according to E 5. 

5. They are disc-like, round, dentated, or serrated bodies (Fig. 

26, page 81). Presence of red corpuscles of the blood. Test 
the urine for coloring matters of the blood with the spectro- 
scope (page 60), also test for hsematin according to Struve, 
page 63. 

6. They are round-like bodies, each having a nucleus. In size 

they are larger than pus corpuscles (Fig. 23, page 79). Pres- 
ence of epithelial cells of the uriniferous tubes of the kidney 



94 CHEMICAL ANALYSIS OF THE URINE. 

or of the urethra of the male. If the cells are somewhat 
numerous they arise from an inflammatory condition of the 
parts. 

Epithelial cells are sometimes granular and contracted, 
and sometimes contain particles of fat, the result of chronic 
disease. 

7. They are large, flat, nucleated bodies (Fig. 24, page 79). Pres- 

ence of epithelial cells of the bladder. 

8. They are round-like bodies with granular contents {a, Fig. 27, 

page 81), which with dilute acetic acid form imperfect nuclei 
{b, Fig. 27). Presence of pus corpuscles (as pus corpuscles 
cannot be distinguished from mucus corpuscles, the presence 
of the former is determined by their number). 

9. They are masses of organized bodies, some cellular and some 

fibrous, with pus and epithelial cells. Presence of patholog- 
ical products. Refer to page 82. 

10. They are exceedingly small bodies, and unless high powers 

of the microscope are employed (1000 diameters), they 
appear as mere points, but in continued movement. In 
fresh urine with high magnifying powers (1200 to 1800 
diameters), the bodies are found to be elongated, formed of 
two or more cells. In urine, three or four days after having 
been passed, they appear as round cells (Fig. 29, page 83). 
Presence of the micrococcus urese. (Refer to Fermentation 
of the Urine, page 20.) 

11. They are cells in numbers of 4, 8, 16, etc. (Fig. 30, page 83). 

Presence of sarcina. 

12. They are filamentous organisms, sometimes found in move- 

ments, each with head and tail but no neck or body (Fig. 28, 
page 81). Presence of spermatozoa. 

13. They are masses of fibres and sometimes accompanied by 

round cells (Fig. 31, page 83). Presence of the ordinary 
mould penicilium glaucum. 



CHAPTER VII. 

Concretions or Stones in the Bladder — Constitution and Physical Properties of Con- 
cretions — Some of the Causes of the Formation of Concretions — Scheme for the 
Analysis of Concretions or Stones. 



CONCRETIONS OR STONES IN THE BLADDER. 
The following constituents of the urine enter into the forma- 
tion of concretions in the bladder, pelvis of the kidneys or ureters. 
Uric acid with the urates, calcium and magnesium phosphates, 
magnesium ammon. phosphate, calcium oxalate, cystin, and xan- 
thin. Of 545 stones, according to Ultzmann, 80.9 per cent, was 
uric acid, S.6 per cent, phosphates, 5.6 per cent, calcium oxalate, 
1.4 per cent, cystin and 3.3. per cent, foreign bodies introduced 
into the bladder or coagulae of blood. Generally, several of these 
bodies are in a concretion. Occasionally, however, either cystin, 
calcium oxalate or ammon. urate is the only constituent. The 
broken surfaces of a concretion formed of different constituents, 
present a stratified appearance, due to layers or strata of the con- 
stituents. Concretions differ very much as regards color, consist- 
ence and other physical properties. Those composed of uric acid 
and urates are hard and dark red in color. Those composed 
chiefly of the phosphates are soft and white. Those composed of 
calcium oxalate are exceedingly hard, white and, when small, 
have smooth surfaces, and as they increase in size their surfaces 
become irregular. If ammon. urate is the principal constituent 
they are soft and light yellow in color. The formation of a con- 
cretion in the bladder is due to crystallization, and the conditions 
favoring the crystallizing process here are the same as they are 
in a beaker glass. Gravel or a sediment formed in the bladder, 
the crystals of which are visible without the aid of a glass, gener- 
ally precedes the formation of concretions, consequently crystals 
or clumps of crystals, or masses of amorphic matter as calcium 
and magnesium phosphates, often become the nuclei of concre- 
tions when by their absence there would be no danger of the 
formation of stones or concretions. Not only do crystals or 
amorphic matter in the bladder often lead to the formation of 

95 



96 CHEMICAL ANALYSIS OF THE URINE. 

stone, but coagulae of blood in the bladder in case of haematuria, 
and foreign bodies introduced into the bladder are known in many 
instances to have caused the formation of stone. It is of great 
importance in cases of gravel, when fresh urine contains free uric 
acid or there is present the phosphates of calcium and magnesium 
due to alkaline reaction of the urine, that the condition of the 
blood be so changed by diet or medication that the urine becomes 
free of gravel or any cloudiness while in the bladder, to avoid one 
of the conditions favoring the formation of concretions. 

QUALITATIVE ANALYSIS OF CONCRETIONS OR STONES. 

(A) After having broken, powder some of the fragments and heat 
a small portion of the powder on a platinum foil or in a 
small porcelain dish over a lamp to dull redness ; continue 
the heat several minutes. It burns completely, leaving no 
residue. 

Presence of cystin, urate of ammonium, uric acid, or xan- 
thin, continue according to A I. It chars or blackens, 
leaving a residue, pass on to B I. 

1. Separate the uric acid, either free or combined, if present, from 

the other constituents by treating some of the powder of the 
stone with an excess of dilute hydrochloric acid in a small 
flask ; warm some time on a water bath. A residue is left. 
Presence of uric acid. Filter while warm, wash with water 
and transfer the residue to a small porcelain dish, treat with 
nitric acid and evaporate to dryness on a water bath. To the 
residue add a small quantity of ammon. hydrate. If a dark 
purple color is produced, it is proof of the presence of uric 
acid. 

2. Evaporate the filtrate of A I to dryness on a water bath and 

mix the residue in a small beaker glass with calcium hydrate; 
add enough water to make a thick paste. Over the beaker 
place a watch glass, to the under surface of which pieces of yel- 
low turmeric and red litmus paper are attached by having 
been moistened with water. At ordinary temperatures, or 
by gently warming the mixture, the yellow paper turns dark 
red or brown, while the red paper turns blue. Presence of 
ammonia. 

3. Treat some of the powder of the concretion with water ren- 



CONCRETIONS IN THE BLADDER. ' 97 

dered strongly alkaline with ammon. hydrate; filter, and to 
the filtrate add a solution of silver nitrate. If a precipitate 
forms, filter and wash with water containing some ammon. 
hydrate, transfer the precipitate to a flask, mix with water 
and separate the silver with sulphureted hydrogen gas. 
Filter and evaporate the filtrate with wash water, in a small 
evaporating dish on a water bath, to dryness. Treat the 
residue with some strong nitric acid, warm gently and add 
a solution of sodium hydrate. A dark red color is evi- 
dence of the presence of xanthin. 
4. Triturate some of the powder of the concretion with water 
rendered strongly alkaline with ammon. hydrate, filter into 
a small beaker glass and wash the residue with water con- 
taining some ammon. hydrate. Evaporate the filtrate with 
wash water in the small beaker over wire gauze to near dry- 
ness. If crystals form, which by microscopic examination 
resemble those represented by Fig. 18, page 76, cystin may 
be present. Dissolve some of the crystals in a solution of 
sodium hydrate and add a small quantity of a solution of 
sodium nitroprusside. If a purple color is produced, cystin 
is known to be present. 

(B ) When some of the powder of the concretion is heated on a 
platinum foil or in a small porcelain crucible, it chars or 
blackens, and leaves a gray-like residue, showing the pres- 
ence of calcium and magnesium phosphates or calcium car- 
bonate from the oxalate. 

1. Separate the uric acid, if present, and test for it according 

to A 1. Preserve the filtrate for testing according to B 
3 and 4. 

2. For xanthin and cystin, separate each from powder of the 

stone and test for each according to A 3 and 4. 

3. Evaporate half of the filtrate of B 1 to dryness on a water 

bath, and examine the residue for ammonia according to 
A 2. 

4. To the other half of the filtrate of B 1 add ammon. hydrate 

to alkaline reaction ; warm, and after the lapse of a few hours 
filter, and wash with water rendered strongly acid with acetic 
acid (to dissolve the phosphates) until the wash water is free 
7 



98 CHEMICAL ANALYSIS OF THE URINE. 

of chlorine, known by producing no precipitate or becoming 
turbid when tested with a solution of silver nitrate, having 
been rendered acid with nitric acid. Dry the filter with 
precipitate and transfer the latter to a small platinum crucible, 
and heat carefully until the bottom of the crucible becomes 
dull red in color. During the application of heat the 
crucible should be covered with a lid. When cold transfer 
the residue in the crucible to a test tube and treat with dilute 
hydrochloric acid, and warm gently. It effervesces. Pres- 
ence of calcium oxalate. By heat the oxalate is changed 
to the carbonate with the liberation of CO, and the carbo- 
nate so formed is decomposed by the acid with the escape 
of C0 2 . By the decomposition of a minute quantity of 
calcium carbonate with hydrochloric acid in a test tube, the 
escape of the gas may not be detected. 

The reaction is made in the field of the microscope by 
placing some of the carbonate on a slide, mixing with a drop 
of water, and when the glass cover is in place, a drop of 
acetic acid is brought to the' margin, and by absorbing water 
from the opposite margin of the cover with a piece of porous 
paper, the acid will come in contact with the carbonate, 
and gas bubbles will be seen evolving in the field of the 
microscope as decomposition takes place. 
5. Incinerate some of the powdered concretion or stone in a 
platinum crucible at a red heat; when cold treat with 
dilute hydrochloric acid; filter, wash the residue, if any, 
and unite the filtrate and wash water containing the cal- 
cium, magnesium and phosphoric acid. Treat the acid 
solution with a solution of sodium carbonate until nearly 
neutral, or slightly acid in reaction, w r hen a solution of ferric 
chloride is added until a drop of the fluid, by means of a 
glass rod brought in contact with some ammon. hydrate in 
a porcelain dish, will produce a buff-colored precipitate. 
The formation of a dirty white precipitate before the 
buff-colored precipitate appears, indicates the presence of 
phosphoric acid. Add to the solution an excess of a solu- 
tion of sodium acetate, heat to the boiling temperature, 
and filter while hot, and wash with water containing 
some sodium acetate. The filtrate contains the calcium and 



CONCRETIONS IN THE BLADDER. 99 

magnesium, which examine according to B 6. Transfer 
the precipitate containing the phosphoric acid, if present, 
to a flask, and boil with a small quantity of a solution of 
sodium hydrate (i part to 10 parts of water). Dilute with 
water and filter. The filtrate contains the phosphoric acid, 
if present, combined with sodium, which is tested with 
magnesia mixture. (A solution of magnesium sulphate treated 
with an excess of ammon. hydrate and the precipitate dis- 
solved with a solution of ammon. chloride.) A white 
crystalline precipitate forms at once, especially if the solu- 
tion contains an excess of ammon. hydrate, or it forms 
after the lapse of a few minutes. Presence of phosphoric 
acid. 

6. The filtrate of B 5 , obtained by filtering after treating with 

ferric chloride and sodium acetate and boiling, is rendered 
distinctly acid with hydrochloric acid, and boiled to drive 
off any carbonic acid that may be present. Test a portion 
of the solution in a test tube with a solution of potassium 
ferrocyanide. It should give no precipitate, showing absence 
of ferric chloride, and when another portion in a test tube is 
rendered alkaline with ammon. hydrate, and an excess of 
a solution of ammon. chloride added, no precipitate should 
be produced, showing absence of phosphoric acid. If ferric 
chloride and phosphoric acid are absent, the filtrate is ren- 
dered distinctly alkaline with ammon. hydrate and an excess 
of a solution of ammon. chloride added. 

The solution is then heated to the boiling temperature 
and a solution of ammon. oxalate added in excess; a white 
precipitate is produced. Presence of calcium. Filter, and 
test the filtrate for magnesium according to B 7. If ammon. 
oxalate produces no precipitate, hence absence of calcium, 
the fluid is tested for magnesium, according to B 7, without 
filtering. 

7. The filtrate from calcium oxalate, B 6, is free of calcium if no 

precipitate forms by the addition of a solution of ammon. 
oxalate ; and if the calcium has been separated, or no calcium 
was found, the filtrate or solution is rendered strongly alkaline 
with ammon. hydrate, and a solution of sodium ammon. 
phosphate added; a white crystalline precipitate is produced. 



100 CHEMICAL ANALYSIS OF THE URINE. 

Presence of magnesium. With the methods here employed 
tests for sulphuric and carbonic acids are not given, neither 
are the tests for potassium and sodium, as they are in mere 
traces in stones or concretions, and therefore their presence 
or absence is not considered in classifying stones or con- 
cretions. 



CHAPTER VIII. 

Filter Paper and Filtering — Evaporating — Drying — Ashing Filters and Heating 
Precipitates — Chemical Balance and Weights — Weighing — Vessels Required for 
Measuring Fluids — Desiccators, Tongs, Crucibles, etc. — Preparation of Solutions 
for Volumetric Analysis — Normal Oxalic Acid — Normal Potassium Hydrate — 
Normal Sulphuric Acid — Normal Hydrochloric Acid — Solution of Litmus — The 
Barium Mixture — The Magnesia Mixture — Millon's Reagent. 



QUANTITATIVE ANALYSIS. 
FILTER PAPER AND FILTERING. 

In quantitative work Schleicher and Schiill's filter paper, No. 
589, is used with advantage, as it leaves no ash by burning. No. 
590 of Schleicher and Schiill is finer paper, and is preferable in 
filtering and washing finely-divided precipitates, as barium sul- 
phate. Either variety of paper is procured, cut in circular pieces. 
The sizes usually employed are 5 j£, 7 and 9 cm. in diameter. 
Swedish filter paper, bearing the name of J. H. Munktell in water 
letters, is the finest filter paper, but it is sold in sheets and leaves 
an ash by burning. To remove the mineral substances from 
Swedish filter paper it is cut in circular pieces by means of a lathe, 
when they are folded in quadrants and placed in a beaker glass 
in order that they may retain their form, and then treated with 
dilute hydrochloric acid — I part acid to 6 parts water. After re- 
maining in contact with the acid a few minutes the latter is drained 
off, and the filters washed with distilled water by standing in con- 
tact with them a few hours and draining off. The process is 
repeated until the last trace of hydrochloric acid disappears from 
the wash water, known by testing with a solution of silver nitrate. 
The filters are then dried by placing them between porous paper 
and kept in a warm place. Swedish filter paper when treated in 
this way is practically free from ash. In regard to the process of 
filtering, funnels having an angle of 6o°, with even surfaces, are 
employed, as the paper Comes in contact with every part of the 
surface and is thereby supported. The margin of the paper, when 
fitted in the funnel, should be about 1 cm. of the brim of the fun- 
nel. Before filtering, the paper is moistened with water and the 

101 



102 CHEMICAL ANALYSIS OF THE URINE. 

filter pressed to the glass, so as to exclude all air bubbles, as their 
presence interferes with the washing. If a fluid composed mostly 
of alcohol is to be filtered, the filter is moistened with alcohol 
instead of water. Each time the fluid is put into the filter, when 
filtering or washing, the amount should not be sufficient to fill 
the filter, as some of the precipitate may pass over the margin of 
the paper and lodge between it and the funnel. At first the clear 
fluid is decanted into the filter, as it passes through the paper 
more rapidly than after the precipitate is brought on the filter. To 
avoid loss, the fluid is introduced from the beaker or dish into the 
filter by means of a stirring rod, placed at the lip of the beaker or 
dish, and the fluid passes down the rod into the filter. In washing 
a precipitate, before each addition of fluid to the filter, any fluid 
already in the filter is allowed to pass through. In transferring the 
precipitate from a beaker glass or dish, a stirring rod, over the end 
of which is placed a short piece of rubber tubing, is employed. 
The rubber will answer the purpose of loosening the precipitate 
from the beaker or dish. To facilitate the process of filtering, 
a filter pump may be employed, or, in the absence of which, 
an aspirator bottle containing water will answer the purpose. 
A platinum cone fitted into the funnel, in which the folded 
end of the filter is placed, will support the paper if the cone 
and paper be properly adjusted. 



EVAPORATING. 

Generally, solutions are concentrated by evaporation on a water 
bath, yet if the fluid contains no bodies mechanically suspended, 
it may be evaporated over the free flame with the heat so regu- 
lated that the fluid will not reach the boiling point. Alcohol and 
ether solutions are evaporated on a water bath at a low tempera- 
ture. In the evaporation of fluids for the purpose of procuring 
crystals for microscopic examination, the solutions should be fil- 
tered, and the process carried on slowly by placing near a stove 
or steam pipes, or if a good air pump is at hand, by removal of 
the atmospheric pressure; especially over concentrated sulphuric 
acid, evaporation will be greatly facilitated. 



QUANTITATIVE ANALYSIS. 103 



DRYING. 



Ill estimating organic substances gravimetrically, as, for 
example, uric acid, kreatinin and albumen, the weight of the 
substance is ascertained by drying the filter paper in an air 
bath at a certain temperature, usually at ioo° C, between two 
watch glasses held in place by a wire spring, and after cooling in 
a desiccator the filter paper, with the watch glasses, is weighed. In 
order that the paper may contain no moisture, the heating and 
weighing are repeated until the weight becomes constant. After 
filtering and washing, the paper and precipitate are dried and 
weighed as before, and the increase in weight is the weight of the 
substance. Bodies that are perfectly insoluble in alcohol and 
ether may be washed with a mixture of alcohol and ether after 
having been washed with water. By this treatment precipitates 
will dry much sooner than by washing with water alone, but in 
exact work the filter should be washed with a mixture of alcohol 
and ether before drying and weighing, as there is some loss in 
weight by the action of alcohol and ether. An air bath is a box 
of sheet iron or copper, provided with a door and shelf or triangle 
support, on which the watch glass or dish containing the substance 
is placed. A thermometer is introduced into the bath through an 
opening in the upper wall. 

ASHING FILTERS AND HEATING PRECIPITATES. 

When the filter and precipitate are thoroughly dry, the latter 
is transferred from the paper to a clean dry watch glass, and the 
paper is folded a few times, and with a platinum wire it is sur- 
rounded by three or four turns, when the filter paper is burned by 
holding it in the oxidizing zones of a Bunsen's flame or spirit 
lamp. Any particles of the precipitate that may have fallen on 
the glazed paper or burner plate over which the operation is con- 
ducted, are collected in the watch glass with a hair pencil. When 
the ash is free. of carbon the platinum wire is held over the pre- 
cipitate and removed from the wire with the hair pencil. If in 
the precipitate there is a reducible metal, as, for example, silver in 
the chloride, loss would likely attend the process, as the metal 
would form an alloy with the platinum wire. Instead of ashing 
on the wire the paper is folded, placed in the weighed crucible 



104 CHEMICAL ANALYSIS OF THE URINE. 

supported by a triangle, and heated gradually until the paper 
ceases to burn with the production of a flame when the crucible 
is heated to redness and usually in fifteen minutes the paper will 
be completely ashed. By inclining the crucible somewhat, air 
will enter in greater quantity. Instead of ashing the paper in 
the crucible the lid of the crucible may be employed for the pur- 
pose, by placing it on the support, with its concave surface upward. 
In case the precipitate is light and small in quantity, it may 
remain with the paper during the process of ashing. To prevent 
loss during the first part of the process, the filter paper with pre- 
cipitate is placed in the crucible closed with the lid, and heat is 
gradually applied until the paper ceases to burn with a flame, 
when the heat is continued with the open crucible until the car- 
bon of the paper disappears by oxidation. Any carbon that may 
have been deposited on the under surface of the lid is removed 
by heating. 

CHEMICAL BALANCE AND WEIGHTS. 

A balance of precision which will bear 100 grammes in each 
pan, and sensitive to o.i milligramme, will answer for all weighing 
required in urinary analysis. The requisites of a good balance 
are given in any modern work on physics. The weights required 
are 50 grms. down to 1 milligrm., with two 10 milligrm. riders. 
Weights, regardless of the reputation of the maker, should be 
tested before using. A set of weights may be too light or too 
heavy, and the results of quantitative work be correct as far as the 
weights are concerned, that is, the 0.5 grm. piece may not be T 5 o 
of I grm. and the 0.05 grm. piece may not be yf-o of 1 grm.; yet, 
if the relative weights are as represented and the weights of the 
set be employed in all weighings, the relative quantities weighed 
would be the same; therefore, in testing a set of weights, standard 
weights, or a standard 5, 10 or 20 grm. piece, need not necessarily 
be employed. However, in preparing some solutions for volu- 
metric work, errors would arise by employing a set of weights 
not absolutely correct, as 50 cc. water measured would not weigh 
50 grms. by the weights. To determine if the weights of a set 
bear correct relationship, place on one pan of the balance, when 
adjusted, the two grm. piece and on the other pan two of the three 
one-grm. pieces, and ascertain if they are of equal weight. Now 



QUANTITATIVE ANALYSIS. 105 

substitute for one of the grm. pieces the third grm. piece and 
determine if they equal in weight the two-grm. piece.. Return the 
weights to the case and place on one of the pans the five-grm.. 
piece, and on the other pan the two and three one-grm. pieces, 
and test as before. The higher weights and the deci- and centi- 
grm. weights, with the centiprm. riders, are tested in the same 
way. In making these tests exact differences are determined by 
employing one of the riders on the arm of the beam with the 
lighter weight. The weights of less than one grm. should not 
differ more than o.i milligrm., and the weights of more than one 
grm. should not differ more than 0.2 milligrm. 



WEIGHING. 

A fine balance of precision should be protected from hydrogen 
sulphide, nitrous fumes and dust. While weighing, every move- 
ment should be guarded that the balance receive no jar ; and when 
the beam is raised from the knife edges or lowered, so as to rest 
on them, the movement should be made with the greatest care, to 
avoid rapid oscillating movements of the beam. The balance is 
in equilibrium when the beam oscillates equally, known by the 
needle leaving the o point an equal number of spaces each oscil- 
lation, and when the beam ceases to oscillate the needle points to 
the o point. The balance is brought in equilibrium by an adjust- 
ment screw at the end of each arm of the beam. When the bal- 
ance is not in use the beam is raised from the knife edges. Usu- 
ally, the body to be weighed is placed on the left pan, but not 
until the beam is raised and the pans arrested. The weights are 
handled with pincers, and under no circumstances with the fin- 
gers, and when the weights are not in use the case containing 
them is closed. The placing of the weights oh the right pan is 
done in order. This will be understood by illustration. Suppose 
the weight of a crucible is 12.5322 grms. The 20-grm. piece is 
found too heavy, next try one of the 10-grm. pieces — too light — 
let it remain and try the 5 -grm. piece — too heavy — return it to the 
case and try the 2-grm.' piece — too light — let it remain and try 
one of the three i-grm. weights — too heavy — return it and try 
the 0.5-grm. platinum piece, and so continue until the additional 
weight required is less than 0.0 10 grm., when the rider is placed 



106 CHEMICAL ANALYSIS OF THE URINE. 

on the beam at number 5, and being found too heavy in this case 
it is moved to 2.5 or 3, and so on until the rider is so located on 
the beam that the latter is in equilibrium. To avoid mistakes in 
recording the weight, first note the weights absent from the case, 
and when they are returned to the case they are again noted, and 
when the numbers are compared, if a mistake has been made, it 
is discovered. In the example given, it is found that the weights 
absent from the case are the 10- and 2-grm. pieces, with the 0.5 
and 0.03-grm platinum pieces, and on the beam the rider is on 
the first mark to the right of number 2 ; hence, the weight is 
12.5322 grms. Bodies are never weighed while warm. 



VESSELS REQUIRED FOR MEASURING FLUIDS. 

A Measuring Flask has a flat bottom and long neck, on the 
lower part of which it is graduated by a mark. They are of 
various sizes, but those generally required are for measuring one 
litre (1000 cc), one-half litre (500 cc), one-fourth litre (250 cc.) 
and 100 cc. 

Glass Cylinders are provided with a base and glass stopper, 
and graduated, the numbers increasing from below upward. 

A cylinder may be employed in measuring any quantity to 
the amount for which it is graduated, that is, with a litre cylinder, 
480, 510, or any other number of cc. within 1000, may be meas- 
ured, but as the diameter of a cylinder is comparatively great, a 
small quantity of fluid making no appreciable difference in read- 
ing the meniscus of the fluid, it cannot be employed where great 
accuracy is required. Cylinders for measuring 100, 250, 500 
and 1000 cc. are usually employed. A cylinder of 2000 cc. is 
convenient for measuring the quantity of urine formed in twenty- 
four hours. 

Pipettes serve the purpose of transferring a definite quantity 
of fluid from one vessel to another. They are of different sizes, 
according to whether they are to measure a definite quantity, for 
example, 5, 10, 20 or 50 cc. for each quantity, employing a pipette 
graduated for the purpose. There are also pipettes, graduated as 
burettes, for measuring from 0.1 cc. to the maximum number of 
cc. for which the pipette is graduated. 

Burettes of 30 and 50 cc. are generally employed. Each 



QUANTITATIVE ANALYSIS. 107 

space representing I cc. is graduated in fifths or tenths. The 
' inferior extremity of a burette is connected with the tip or 
dropper by a piece of vulcanized rubber tube, which is secured 
by a Mohr's pinch-cock, to regulate the quantity of fluid in titra- 
ting. This piece of apparatus is preferable to a glass stop-cock, 
but a short piece of glass rod, with its ends rounded by heating 
in the flame of a blowpipe, placed in the rubber tube will answer 
every purpose, if it fits the tube close enough to prevent the 
passage of fluid. By pressing the sides of the rubber tube with 
the finger and thumb it will separate from the plug, unless the 
latter be too large, and the fluid will pass through the openings. 
If, after a burette or pipette has been washed with water, and 
dried, drops of the fluid it is intended to measure adhere to its 
sides, it should be well washed with akohol and ether before 
using. 

The burette is filled by placing the tip, or dropper, in the fluid, 
and while the pinch-cock is loosened by pressure, the fluid is 
drawn up by suction in quantity sufficient to fill the flower end 
of the burette, when the rubber tube is secured, and a small 
funnel is placed in the upper end of the burette, and the fluid is 
carefully poured into the burette until it stands but a short dis- 
tance above the o mark. The fluid is now allowed to pass out 
of the burette drop by drop, by pressing the pinch-cock until the 
meniscus of the fluid coincides with the o mark. By the use of 
Erdmann's float readings are made with greater precision. In 
reading the meniscus of a fluid in a burette, sufficient time 
should elapse before the reading, so that the fluid adhering to the 
surface may run down; the eye should be on the same plane as 
the meniscus of the fluid, and all readings should be conducted 
on the same basis; that is, if the background is light, and the 
dark line below the light segment is taken as the meniscus in the 
first reading, it should be taken in all subsequent readings. 

Measuring vessels, whether flasks, pipettes or burettes, should 
be tested before using. They are graduated to measure fluids at 
17.5 ° C. As 1 cc. distilled water at this temperature weighs I 
grm., pipettes and burettes are tested by weighing a measured 
quantity of distilled water on a balance of precision. For this 
purpose, balance a beaker glass with a watch glass containing 
shot and tin foil, and into the beaker introduce a measured quan- 



108 CHEMICAL ANALYSIS OF THE URINE. 

tity of water at 17.5 . In testing burettes a weighing is made for 
every 10 cc. If in weighing 10 cc. water the weight comes 
within 10 milligrms. of 10 grins., the burette may, according to 
Fresenius, be considered correctly graduated. If, however, an 
Erdmann's float be employed, the difference should not exceed 2 
milligrms. In testing graduated flasks of more than 100 cc. a 
balance which will bear the increased weight and show a differ- 
ence of 0.1 milligrm. is required. 

DESICCATORS, TONGS, CRUCIBLES, ETC. 

Of the various forms of desiccators that of Fresenius, having a 
capacity of 1 500 cc, is preferable, as a crucible is placed in or taken 
out by means of the tongs without obstruction. By the application 
of some grease to the ground surfaces of the lid, and employing 
concentrated sulphuric acid as the dryer, the apparatus answers 
the purpose. Brass tongs are kept clean without much trouble, 
and do not rust or corrode, as iron. They should be bent so as 
to transfer crucibles without inconvenience. 

The Royal Berlin Porcelain Crucibles are the best quality, being 
thin, light, well glazed, and resist sudden change of temperature 
without breaking. Those having a capacity of 14 cc. are conve- 
nient for ashing filter paper, heating precipitates, and weighing. 

A platinum crucible of 20 or 22 grms., including weight of 
cover, and capacity of 22 cc, will answer every purpose in urinary 
analysis when a platinum crucible is preferred. A platinum dish, 
with cover, weighing 40 to 45 grms., having a capacity of 50 cc, 
is a convenient size for estimating the chlorides of potassium and 
sodium. 

Other articles required, but not enumerated here, are : a burner 
plate, over which to burn filters; a piece of platinum wire, No. 5, 
on which to burn filters ; a sheet of glazed paper, a pair of small 
pincers and a fine hair brush or pencil. 

PREPARATION OF SOLUTIONS FOR VOLUMETRIC ANALYSIS. 

The system of standardizing solutions employed in volumetric 
analysis is based on the valence of one atom of hydrogen. Of a 
monobasic acid, its molecular weight in grammes occupying the 
volume of 1000 cc is normal. The molecular weight of nitric 



QUANTITATIVE ANALYSIS. 109 

acid, HN0 3 , is 6$. Therefore, 63 grms. pure nitric acid diluted 
with water to 1000 cc. is normal acid. The molecular weight of 
potassium hydrate, KOH, is 56; hence, 56 grms. in 1000 cc. is 
normal potassium hydrate. The molecular weight of sodium 
hydrate is 40 ; hence, 40 grms. in 1 litre. It is understood that 
one-half of the molecular weight of a dihydric acid or a dibasic 
oxide in grammes, dissolved in water and diluted to the volume 
of 1000 cc, has equal chemical value to the full molecular weight 
of a monohydric acid or monobasic hydrate in grammes occupy- 
ing the volume of 1000 cc, In case a compound contains water 
of crystallization, the molecular weight of the water is added to 
that of the compound ; for example, oxalic acid has the formula — 

C 2 H 2 4 ,2H 2 0. 

The molecular weight of the acid proper, C 2 H 2 4 , is 90, and that 
of the water, 2H 2 0, is 36. 90 + 36 == 126. 126 is therefore the 
molecular weight of the acid, and as this acid is dibasic, y 2 of 126 
in grammes, 63, occupying the volume of 1000 cc, is normal. In 
standardizing solutions of alkalies and acids, pure oxalic acid may 
be employed. To avoid error, however, the acid marked chem- 
ically pure is dissolved in hot water to saturation, filtered while 
hot, and the filtrate stirred with rapidity in a beaker surrounded 
by cold water until the temperature is reduced. Fine crystals 
will separate. Filter, and wash the crystals on the filter with a 
small quantity of cold water and place between porous paper. 
Commercial oxalic acid is purified by recrystallizing several 
times. 

NORMAL OXALIC ACID (63 grms. C 2 H 2 4 ,2H 2 in 1000 cc. WATER). 

Pure oxalic acid is placed between filter paper and pressed 
until dry, when about 20 grms. are weighed on a balance of preci- 
sion and introduced into a clean, dry flask. The quantity of water 
in which to dissolve the acid is determined by the equation 
63 : 1000 : : weight of oxalic acid : x. x = No. of cc. water. 
The solution will undergo no change if heated to near the boiling 
point in a flask which is closed with a rubber stopper while the 
solution is hot, and kept in a cool, dark place. 

NORMAL POTASSIUM HYDRATE (56 grms. KOH in 1000 cc. WATER). 

Potassium hydrate is preferred to sodium hydrate on account 
of the tendency of the latter to etch the burettes. As either com- 



110 



CHEMICAL ANALYSIS OF THE URINE. 



pound " chetn. pure" usually contains more or less of the car- 
bonate, the latter is separated by treating a solution with barium 
hydrate until after mixing well by shaking a small quantity, fil- 
tered into a test tube containing a solution of barium hydrate, will 
produce no cloudiness. Another portion is filtered, and the filtrate 
tested with dilute sulphuric acid, and if a light precipitate or 
cloudiness is produced, barium hydrate has been added to the 
solution of potassium hydrate in excess, to separate which add 
small quantities of a solution of potassium carbonate, mixing well 
by shaking after each addition. The solution is thus treated with 
barium hydrate and potassium carbonate until small portions, 
filtered, will produce no cloudiness with either barium hydrate or 
dilute sulphuric acid. A solution of potassium hydrate or sodium 
hydrate may be prepared by dissolving yo grms. of the former or 
50 grms. of the latter in 1 100 cc. water; and after separating the 
carbonic acid, titrating with the normal oxalic acid as below, or a 
larger quantity of a much stronger solution may be prepared ; 
and having separated the carbonic acid from the solution (employ- 
ing a solution of sodium carbonate to separate excess of barium 
hydrate from the sodium hydrate solution), when the solution is 
diluted for titration, by first determining its specific gravity with a 
hydrometer from which the percent, of either KOH or NaOH is 
ascertained by consulting Table 2 or 3. 



TABLE 2.— SHOWING PERCENTAGE OF KOH IN AQUEOUS SOLUTIONS CORRE- 
SPONDING TO THE SPECIFIC GRAY., TEMP. i 5 °. {LUNGE.) 



SPECIFIC 
GRAVITY. 



PERCENTAGE. 



I.060 
I.067 
I-075 
I 083 
1. 091 
I. IOO 
1. 108 
I.II6 
1. 125 



7-4 
8.2 
9.2 



10.9 
12.0 
12.9 
13-8 
14.8 
I 5-7 



SPECIFIC 
GRAVITY. 


PERCENTAGE. 


1. 142 


| 
16.5 


1. 152 


17.6 


1. 162 


18.6 


I.I7I 


19.5 


1. 180 


20.5 


I.IQO 


21.4 


I.200 


22 4 


1. 2IO 


23-3 


I.220 


24.2 


I-23I 


25-1 



PERCENTAGE. 



26.I 
27.O 
28.O 
28.9 
29.8 
3°-7 

3i-8 

32-7 

33-7 
34-9 



QUANTITATIVE ANALYSIS. 



Ill 



TABLE 3.-SHOWING PERCENTAGE OF NaOH IN AQUEOUS SOLUTIONS CORRE- 
SPONDING TO THE SPECIFIC GRAY., TEMP. 15° {LUNGE.) 



SPECIFIC 
GRAVITY. 


PERCENTAGE. 


SPECIFIC 
GRAVITY. 


1 
PERCENTAGE. 


SPECIFIC 
GRAVITY. 


PERCENTAGE. 


I.075 


6 55 


1. 162 


J 4-37 


I.263 


23 


67 


I.083 


7-31 


1. 171 


15-13 


I.274 


24 


81 


1. 091 


S.OO 


1. 180 


15-91 


,,.8 5 


25 


80 


I. TOO 


8.68 


1. 190 


16.77 


I.297 


26 


83 


1. 108 


9.42 


I.200 


17.67 


I.308 


27 


80 


1.1x6 


10.06 


1. 2IO 


18.58 


I.320 


28 


83 


1. 125 


10.97 


I.220 


19.58 


1-332 


29 


93 


1-134 


11.84 


1. 231 


20.59 


1-345 


31 


22 


1. 142 


12.64 


1. 241 


21.42 


1-357 


32 


47 


1. 152 


13-55 


I.252 


22.64 


1-37° 


33 


69 



If, for example, the specific gravity of a solution of potassium 
hydrate is 1.22, the per cent, of KOH is 24.2; therefore, in 1 cc. 
weighing 1.22 grm. there is 0.29524 grm. KOH ( 24 " 2 * 1-22 = 
0.29524), and in 1000 cc. normal potassium hydrate there being 56 
grms. KOH, this quantity is in 189.6 cc. of the solution having 
the sp. gr. 1.22 (^^ = 189.6). But the method of determin- 
ing the quantity from the specific gravity of a solution is not 
accurate, otherwise the normal solution could be prepared by 
diluting 189.6 cc. of the solution with water to 1000 cc. To 
titrate with normal oxalic acid, the solution is diluted so that 
the solution be somewhat stronger than that of normal strength. 
For this purpose 230 cc. of the solution is diluted to 1100 cc. 
Having mixed well by shaking, the solution is standardized 
with normal oxalic acid. 

Introduce 20 cc. of the solution of potassium hydrate from 
a burette into a 400 cc. flask, add 100 cc. water and sufficient 
solution of litmus (the preparation of which is found below) 
to impart a distinct, but not deep blue, color to the solution. 
Titrate the solution with normal oxalic acid until, after mixing 
by agitating the flask, the color of the solution turns purple or 
red by the addition of 0.1 cc. Repeat the titrations until an 
agreement is reached, when the solution is diluted so that 20 cc. 



112 CHEMICAL ANALYSIS OF THE URINE. 

will correspond to 20 cc. of the normal oxalic acid. If, for exam- 
ple, 22.2 of the normal acid is required to neutralize 20 cc. of the 
solution of potassium hydrate to dilute the latter to normal 
strength, 20 cc would require the addition of 2.2 cc. water and 
1000 cc. would require no cc. water (20 : 2.2 : : 1000 : x). To 
prevent a standardized alkaline solution, whether of potassium, 
sodium or barium hydrate, from absorbing carbonic acid of the 
air, the bottle containing the solution is provided with a rubber 
stopper having a hole into which a glass tube is introduced, which 
is connected with a U-shaped tube containing fragments of soda 
lime. 

NORMAL SULPHURIC ACID (49 grms. H 2 S0 4 in icoo cc. WATER). 

As pure concentrated sulphuric acid always contains I to 4 per 
cent, water, the normal acid is not prepared by weighing the con- 
centrated acid on a balance of precision and dissolving in water. 
Either 60 grms. of the concentrated acid is diluted with water to 
1 100 cc. or the specific gravity of dilute sulphuric acid is deter- 
mined with a hydrometer, and the per cent, of H 2 S0 4 ascertained 
by consulting Table 4, and a definite quantity of the acid diluted, 
so that the solution be somewhat stronger than normal, when it 
is titrated with normal potassium hydrate. For example, the spe- 
cific gravity of dilute sulphuric acid is 1,20, the per cent, of 
H 2 S0 4 is 27.1, I cc. of the acid would therefore weigh 1.2 grm. 
and contain 0.3252 grms. H 2 S0 4 ( I-2 *J 7 ' Z = 0.3252). As 1100 
cc. of normal sulphuric acid contain 53.9 grms. H 2 S0 4 , so 165.7 
cc. of the dilute acid contain about this quantity of H 2 S0 4 
(^^ = 165.7), Dut tne solution is prepared stronger for titration, 
therefore 175 cc. of the dilute acid is diluted with water to 1100 
cc. Introduce 20 cc. normal potassium hydrate from a burette 
into a 400 cc. flask, and having added about 100 cc. water and 
solution of litmus (the preparation of which is found below), 
to impart a blue color to the solution, but avoiding an excess 
titrate with the sulphuric acid — diluted — until, after mixing the 
solution by agitating the flask, the color turns purple or red by 
the addition of 0.1 cc Repeat the process until an agreement is 
reached, when the acid is diluted to normal strength. If 18.8 cc. 
of the acid be required to neutralize 20 cc. of the normal potas- 
sium hydrate, this quantity of acid would require 1.2 cc. water 



QUANTITATIVE ANALYSIS. 



113 



to become normal, and iooo cc. would require 63.8 cc. water 
(18.8 : 1.2 : : 1000 : x. x = 63.8). 



TABLE 4.— SHOWING PERCENTAGE OF H 2 S0 4 IN DILUTE ACIDS CORRESPOND- 
ING TO THE SPECIFIC GRAVITY, TEMP. i 5 ° C. {KOLB.) 



SPECIFIC 
GRAVITY. 


PERCENTAGE. 


SPECIFIC 
GRAVITY. 


PERCENTAGE. 


SPECIFIC 
GRAVITY. 


PERCENTAGE. 


I.060 


8.8 


1. 142 


19,6 


I.24I 


32.2 


I.067 


98 


1.152 


20.8 


I.252 


33-4 


I.075 


IO.S 


1. 162 


22.2 


1.263 


34-7 


I.083 


11.9 


I.I7I 


23-3 


I.274 


36.0 


I.09I 


13 00 


1. 180 


24 5 


I.2S5 


37-4 


I. IOO 


14. 1 


1. 190 


25.3 


1.297 


38.8 


1. 108 


15-2 


I.200 


27 1 


I.308 


40.2 


I. Il6 


16.2 


1. 2IO 


28.4 


I.320 


41.6 


1. 125 


17-3 


I.220 


29.6 


I 332 


43° 


i-i34 


18.5 


I.23I 


31.0 


1-345 


44-4 



NORMAL HYDROCHLORIC ACID (36.46 grms. HC1 in 1000 cc. WATER). 

Mix 190 cc. pure hydrochloric acid, specific gravity 1.12, with 
about 900 cc. distilled water, or determine the specific gravity of 
dilute acid, and, from the specific gravity, having ascertained the 
per cent, of HC1 by consulting Table 5, the weight of HC1 in 1 
cc. of the acid is divided into 36.46; the quotient is about the 
number of cc. of the acid required for 1000 cc. of the normal 
acid. For example, the specific gravity is 1.166, the per cent, of 
HC1 is 33. 1 cc. of the acid weighs 1.1 66 grms. and contains 



038478 grm. HC1 (- 



[66 X 33 



0.38478). As i cc. of the dilute 



acid contains 0.38478 grm. HC1, and in 1000 cc. normal acid there 
are 36.46 grms. HC1, as many cc. of the dilute acid contains the 
required quantity of HC1 as 0.38478 is contained in 36.46, which 
is 94.7; therefore, 94.7 cc. of the acid diluted with water to 1000 
cc, or 104.2 cc. diluted- to 1100 cc, would be about normal 
strength ; but the acid is prepared for standardizing by titrating 
with normal potassium hydrate somewhat stronger; hence 120 
cc. is 8 diluted with water to 1100 cc. 



114 CHEMICAL ANALYSIS OF THE URINE. 

Introduce 2D cc. normal potassium hydrate, page 109, into a 
400 cc. flask, add about 100 cc. water and a sufficient quantity 
of solution of litmus (refer below) to impart a distinct blue 
color to the fluid, but avoid the addition of any more litmus than 
necessary. Titrate the solution with the acid (diluted) from a 
burette until, after agitating the flask, the color of the solution 
becomes purple or red by the addition of 0.1 cc. Repeat the 
titrations until an agreement is reached when the acid is diluted 
to normal strength. If 20 cc. normal potassium hydrate requires 
1 8. 1 cc. of the acid, the addition of 1.9 cc. water to this quantity 
of acid would be required, and 1000 cc. would require dilution 
to 1 104.9 cc - (18.1 : 1.9 : : 1000 : x. x = 104.9). 



TABLE 5.— SHOWING PERCENTAGE OF HCL IN DILUTE ACID CORRESPOND- 
ING TO SPECIFIC GRAVITY, TEMP. i 5 c C {KOLB.) 



SPECIFIC 
GRAVITY. 


PERCENTAGE. 


SPECIFIC 
GRAVITY. 


PERCENTAGE. 


SPECIFIC 
GRAVITY. 


PERCENTAGE. 


I.067 


*3-4 


I.I34 


26.6 


I. l8o 


35-7 


I.075 


15.0 


I-I43 


28.4 


1. 185 


36.8 


I.0S3 


16.5 


1. 152 


30.2 


1. 190 


37-9 


1. 09I 


18.1 


1. 157 


31.2 


I-I95 


39-° 


I IOO 


19.9 


1. 161 


32.O 


1. 199 


39-3 


1. 108 


«.s 


1. 166 


33-o 


I.205 


41.2 


I. Il6 


23.1 


1. 171 


33-9 


1. 2IO 


42.4 


1. 125 


24.8 


I.I75 


34-7 


1. 212 


42.9 



SOLUTION OF LITMUS. 
Pulverize 50 grms. litmus and introduce the powder into # a 
flask containing 300 cc. distilled water, warm one or two hours 
on a water bath, frequently shaking, and decant through a filter. 
Divide the filtrate into two equal volumes, and by means of a 
glass rod introduce small quantities of either dilute nitric or sul- 
phuric acid into one portion of the solution until the color 
becomes purple or red, avoiding the addition of more acid than 
necessary, when the two volumes are united and 50 cc. strong 
alcohol added. The solution is preserved by keeping in a dark, 



QUANTITATIVE ANALYSIS. 115 

cool place in small bottles filled to the neck, each closed with a 
cork having a groove cut in one side to permit access of air. 

THE BARIUM MIXTURE. 

The barium mixture employed in urinary analysis is composed 
of saturated solutions of barium hydrate and nitrate. It is pre- 
pared by dissolving about ioo grms. pure crystallized barium 
hydrate in iooo cc. warm water, and when cold, if all the hydrate 
is dissolved, more is added, and the solution is again well mixed 
by shaking. The saturated solution of barium nitrate is pre- 
pared in the same way, when two volumes of the solution of 
barium hydrate are mixed with one volume of the solution of 
barium nitrate. The mixture is kept in well-stoppered bottles. 

THE MAGNESIUM MIXTURE. 

The magnesia mixture employed in separating phosphoric 
acid from solution, is prepared by dissolving 56 grms. magne- 
sium chloride in 400 cc. distilled water in a 1000 cc. graduated 
flask, and adding 70 grms. ammon. chloride, and when solution 
has taken place, 350 cc. strong ammon. hydrate is added, and the 
flask is filled with water to the mark. 

MILLON'S REAGENT. 

Dissolve one part of mercury by weight in one part of concen- 
trated nitric acid by weight. For this purpose a flask may be 
employed. Usually*, the mercury dissolves in a few minutes, 
when the solution is diluted with an equal volume of water. 



CHAPTER IX. 

The Quantity of Urine Passed in Twenty-four Hours — Specific Gravity — The Solids 
— Inorganic Substances — The Coloring Matter — Acidity of the Urine — Urea — 
Liebig's Method Modified by Pfliiger — Estimation of Urea in Diseased Urine by 
Liebig's Method — Knop's Method Modified by Greene. 



THE QUANTITY OF URINE PASSED IN TWENTY-FOUR HOURS. 

For the purpose of determining the quantity of urine formed 
in twenty-four hours, a graduated cylinder of 2000 cc., divided in 
spaces of 10 cc, is employed, but not having a cylinder so large, 
one of 500 or 1000 cc. will answer the purpose by emptying and 
refilling, taking care, however, to drain well before refilling. The 
result of the quantitative estimation of sugar, albumen, etc., in 
urine passed, for example, in the morning, is of little value, as the 
quantity of urine formed during one period varies according to 
the quantity of fluids taken. The result is of value when it 
embraces the quantity in urine formed in periods of twenty-four 
hours. Therefore, the determination of the quantity of urine 
passed during this period is nearly always the first step in all 
quantitative estimations of constituents of the urine, whether of 
normal or diseased urine. 

In order that the calculation of the quantity of a constituent of 
the urine be made simple, round numbers in cc. are taken ; for 
example, if 1560 cc. is the total quantity of urine of twenty-four 
hours, the number is made 1600. The slight difference does not 
practically change the result. 

SPECIFIC GRAVITY. 

There are three methods for determining the specific gravity of 
the urine, by means of a urinometer, a Mohr-Westphal's balance, 
and a picnometer. As the results obtained by the use of the 
urinometer are sufficiently accurate, except in estimating sugar 
by the fermentation method, it is seldom that the other methods 
are employed. For the conditions for the employment of the 
urinometer, refer to Specific Gravity, Chapter vi. 

116 



INORGANIC SUBSTANCES. 117 

THE SOLIDS OF THE URINE. 

By evaporating a definite quantity of urine on a water bath to 
dryness, and subjecting the residue to the temperature of ioo° C. 
a few hours and weighing the residue, the result is not correct, 
as some of the urea decomposes during the process of evapora- 
tion. The body that escapes is ammonia, and by the absorption 
and estimation of which accurate results are obtained. However, 
with less work, and with results equally as accurate, a small quan- 
tity of urine, 5 cc, is evaporated in a platinum dish over strong 
sulphuric acid in vacuo, the air having been removed by means 
of an air pump. When it is found that by renewing the acid 
the weight is constant, subtract the weight of the dish from the 
weight of the dish -f- contents, the difference is the weight of 
the solids in 5 cc. urine, and by multiplying by 20 the product is 
the number of parts in 100 parts of the urine. It has been found 
that by multiplying the number above 1000 which the urine 
weighs more than distilled water by 2.33 (Hasser's number) the 
product expresses the number of parts of solid matter in 1000 
parts of the urine; for example, urine having specific gravity of 
1020. 20 X 2.33 = 46.6 parts solids in 1000 parts urine. The 
average total quantity of solid matter in the urine of twenty-four 
hours of an adult is about 60 grms. — between 4 and 5 per cent. 

INORGANIC SUBSTANCES. 

In a weighed platinum dish evaporate 50 cc. clear or filtered 
urine on a water bath to dryness, heat the dish carefully over a 
spirit lamp or Bunsen's burner by keeping the flame, which 
should not be large, in continued movement, so that all parts of 
the dish are heated equally. When vapors are no longer given 
off, remove the lamp, and, when cold, treat the charred mass with 
some hot distilled water; stir with a glass rod and filter through 
a small filter paper into a clean flask of about 200 cc. capacity. 
This process is repeated until the wash water is free of the chlo- 
rides, known by forming no cloudiness, when a small quantity is 
tested with a solution of silver nitrate and some nitric acid. The 
evaporating dish containing some of the charred mass is then 
placed on a water bath, and when the contents are dry, it is 
gradually heated to a dull, red color, when the carbon will gradu- 



118 CHEMICAL ANALYSIS OF THE URINE. 

ally oxidize and a gray ash be left. The filter paper, which may 
contain some of the insoluble constituents, is dried, ashed in the 
dish, when the filtrate with wash water in the flask is evaporated 
in the dish on a water bath to dryness. Care is taken that there 
is no loss of the filtrate or wash water, and that the flask contain- 
ing them is rinsed several times with distilled water and the rins- 
ings evaporated in the dish. The dish is then heated carefully to 
dull redness, placed in a desiccator, and, when cold, it is weighed. 
The difference in weight between that of the empty dish and that 
of the dish -f- contents is the weight of inorganic matter in 
50 cc. urine, and by multiplying by two the product is the weight 
of inorganic matter in 100 cc. urine. If, instead of filtering 
and evaporating the filtrate, etc., the urine is evaporated to dry- 
ness and the residue ashed, part of the chlorides would pass off, 
while part would fuse and prevent complete oxidation, hence, the 
result would be incorrect. To avoid these sources of error the 
method is more lengthy than it otherwise would be. 

THE COLORING MATTER. 

There is no method for estimating the quantity of urobilin in 
the urine; besides, it is not the only coloring matter in urine, as 
shown by the spectroscope. 

THE ACIDITY OF THE URINE. 

The degree of acidity of the urine is determined by titrating 
with a standardized solution of potassium or sodium hydrate. 
The T \j normal is preferable to either the i or the normal solu- 
tion. For the preparation of normal potassium hydrate, refer to 
page 109. From the normal solution the ■£$ is prepared by intro- 
ducing 100 cc. of the solution into a litre (1000 cc.) flask and 
filling with distilled water to the mark. After mixing well by 
shaking, the solution is ready for use. In titrating, litmus paper 
is used as the indicator. The estimation is made by filling a 100 
cc. pipette to the mark with the urine, and having introduced it 
into a beaker glass it is titrated with the T Vj normal potassium 
hydrate from a 25 or 30 cc. burette. The burette, before filling 
with the solution, should be clean and dry. In titrating, the urine 
is frequently tested with red litmus paper, and when the paper 



UREA. 119 

changes to purple or blue by the addition of one or two drops of 
the solution, after stirring well with a glass rod, a sufficient quan- 
tity of potassium hydrate has been added, and after standing a 
short time the meniscus of the fluid in the burette is read. Titra- 
tions are repeated until an agreement is reached. The degree of 
acidity of the urine may be calculated as quantity of oxalic acid. 
For example, if ioo cc. urine required 20 cc. of the ys normal 
solution of potassium hydrate, the quantity of oxalic acid in 20 cc. 
of the tV normal solution (J--$ of the quantity in 1000 cc.) is 0.126 
grm. ; hence, the acid salts in 100 cc. of the urine equal in satu- 
rating effect 0.126 grm. oxalic acid. Suppose the quantity of 
urine formed in twenty-four hours is 1600 cc, then 16 X 0.126 
= 2.016 grms. oxalic acid. 

UREA. 

Notwithstanding a great many methods for the estimation of 
urea in the urine have been proposed, Liebig's method, as modi- 
fied by Pfliiger and others, and the method of Knop and Greene, 
may properly be regarded as the most trustworthy. 

LIEBIG'S METHOD AS MODIFIED BY PFLUGER. 

This method is based on the fact that when a solution of mer- 
curic nitrate comes in contact with a solution of urea, a precipi- 
tate forms composed of two bodies — 

2CH,ON 2 , Hg(N0 3 ) 2 , 3 HgO. 

However, in the reaction there is some nitric acid set free — 

(2CH 4 ON 2 + 4 Hg(N0 3 ) 2 + 3H 2 = (CH 4 ON 2 ) 2 Hg(N0 3 ) 2 , 3 HgO + 6HNO s ), 

which dissolves more or less of the urea compound and pro- 
duces changes in its constitution, consequently the acid is nearly 
neutralized while titrating. When a certain excess of a solution 
of mercuric nitrate has been added to a solution of urea, and the 
fluid is brought in contact with a solution of sodium bicarbonate, 
a yellow precipitate forms ; therefore, in titrating, sodium bicarbo- 
nate is employed to determine when the urea has combined with 
the mercury and an excess of the mercuric nitrate is in the solu- 
tion. It was found that a solution of the mercury salt to form 
the yellow compound with sodium bicarbonate contains 3.47 
milligrms. mercuric oxide in ice. From this it is readily seen 



120 CHEMICAL ANALYSIS OF THE URINE. 

that results vary according as dilute or strong solutions of urea 
are employed, for the excess (3.47 milligrms. HgO in 1 cc.) is the 
same in either case. In standardizing the mercury solution with 
a two per cent, solution of urea, the former is diluted so that 20 
cc. corresponds to 10 cc. of the solution of urea; therefore, in 
titrating, the mixture occupies a volume of 30 cc, when the yel- 
low coloration or precipitate takes place by bringing some of 
the mixture with sodium bicarbonate. The solution or mixture, 
therefore, contains 104.1 milligrms. HgO, more than is sufficient 
to combine with the urea (30 X 3.47 — 104. 1). Now, suppose 
that instead of the 2 per cent, solution of urea a 1 per cent, 
solution be employed, or the 2 per cent, solution be diluted with 
an equal volume of water and 20 cc. be titrated ; in this case the 
quantity of urea present is the same, but there is 10 cc. more 
water. When 20 cc. of the mercuric nitrate solution is added, 
the yellow precipitate will not appear when some of the mixture 
is brought in contact with sodium bicarbonate, for the reason 
that 1 cc. of the mixture contains but 2.6 milligrms. HgO instead 
of the quantity required, 3.47 milligrms. (^^ = 2.6). To the 
mixture (occupying the volume of 40 cc. instead of 30 cc.) there 
is required the addition of 34.7 milligrms. HgO, that there may 
be present the excess of HgO, required to yield a yellow precipi- 
tate with sodium bicarbonate. Without correction, the addi- 
tional quantity, 34.7 milligrms. HgO, would be accredited to urea 
that is not present; hence, estimations of urea are too high by 
employing dilute solutions, or solutions of less than 2 per cent, 
urea. And in the employment of solutions of greater strength 
than 2 per cent., the excess of HgO required, in proportion to the 
amount of urea, is less, consequently the estimations are too low. 
The phosphoric acid of the phosphates in the urine combines 
with the mercury of the nitrate, so that its removal from the 
urine before making the titration is requisite. The presence of 
the chlorine of the chlorides in the urine also interferes with the 
reaction between the mercuric nitrate and urea, as mercuric 
chloride is formed which does not combine with urea, neither 
does it yield the yellow precipitate with sodium bicarbonate ; 
consequently, in the presence of the chlorides more mercuric 
nitrate is required, and its removal from the urine before making 
the estimation is necessary. 



SOLUTION OF SODIUM CARBONATE. 121. 

PREPARATION OF SOLUTIONS REQUIRED IN LIEBIG— PFLUGER'S METHOD. 
SOLUTION OF UREA. 

Having dried some pure urea by remaining in a watch glass 
over concentrated sulphuric acid in a desiccator, until by 
repeatedly weighing there ceases to be any further loss of 
weight, 2 to 4 grms. is weighed and introduced into a clean, dry 
flask of 250 or 400 cc. capacity. To the urea add the required 
amount of distilled water in cubic centimetres, to standardize to 
the strength of 2 grms. to 100 cc. ; therefore, I cc. of the solution 
would contain 0.020 grm. urea, or the solution is 2 per cent. urea. 

THE SOLUTION OF MERCURIC NITRATE. 

About 87 grms. of the yellow mercuric oxide is put into a por- 
celain dish, and treated with one volume of nitric acid, sp. gr. 
1.20, and one volume of water. Heat on a water bath, and when 
brown fumes cease to evolve, add portions of the dilute acid 
until by heating brown fumes are no longer given off. Evapo- 
rate to syrupy consistence while it is stirred, until acid vapors 
cease to evolve. The mercuric nitrate so formed is dissolved in 
1 100 cc. distilled water, but to avoid the formation of the basic 
nitrate in preparing the solution, water is added to the salt in 
small quantities, while the dissolving process is facilitated by stir 
ring, when the solution is carefully poured into a graduated 
glass cylinder of 1200 or 1500 cc. capacity, and any undissolved 
residue in the dish is dissolved in a small quantity of the 
dilute acid, and the solution is transferred to the cylinder, 
and, finally, the dish is rinsed with distilled water and the rinsings 
added to the contents of the cylinder. The solution in the 
cylinder is diluted by adding water in small quantities, fol- 
lowed by shaking so as to mix well, until the fluid reaches the 
1 1 00 cc. mark. Having mixed well by shaking, the solution is 
ready for standardizing with the solution of urea. 

THE SOLUTION OF SODIUM CARBONATE. 

Having heated pure sodium carbonate in a dish over a lamp to 
dryness, 53 grms. is introduced into a litre flask and with distilled, 
water filled to the mark. In this case the weight of the salt may 
be approximative. 

For the preparation of the barium mixture, refer to Chapter vin. 



122 CHEMICAL ANALYSIS OF THE URINE. 

SOLUTION OF SILVER NITRATE. 

Dissolve pure crystallized silver nitrate in distilled water, so 
that the strength of the solution will be of 29.075 grms. to one 
litre of water. Refer to Volhard's method for the estimation of 
chlorine, Chapter xi. 

TO STANDARDIZE THE SOLUTION OF MERCURIC NITRATE. 

The solution of mercuric nitrate as prepared above is too 
strong, and to determine what dilution is required it is titrated 
with the urea solution. For this purpose, introduce 10 cc. of the 
urea solution by means of a clean, dry pipette into a small 
beaker. A 25 or 30 cc. burette is cleaned, dried and filled to the 
o mark with the mercuric nitrate solution, and 19 cc. is intro- 
duced without interruption into the beaker with the urea solu- 
tion, when the mixture is stirred with a glass rod, and titrated 
with the solution of sodium carbonate from a burette, until the 
mixture is nearly neutral. 0.2 or 0.3 cc. of the mercury solution 
is now added, and, after stirring, transfer a drop of the mixture 
by means of a glass rod to a small quantity of a mixture of 
sodium bicarbonate and water in a watch glass, placed on a black 
background, and if the precipitate formed is white add another 
0.1 cc. of the mercury solution, and having mixed well by stir- 
ring, if a drop of the mixture produces a yellow coloration, 
or precipitate, with sodium bicarbonate, enough of the mercury 
solution has been added. If the precipitate is still white, and 
since neutralizing 0.4 or 0.5 cc. of the mercury solution has been 
added, the excess of acid is again nearly neutralized, when the 
mercury solution is added in quantities of 0.1 or 0.2 cc. until a 
drop of the mixture yields a yellow coloration with sodium 
bicarbonate. In making the second titration, introduce without 
interruption within 0.2 cc. of as much of the solution as was 
required in the first titration; stir well, titrate with the soda solu- 
tion until nearly neutral, and, having added 0.1 cc. of the mer- 
cury solution, test as before, and repeat the process until a yellow 
coloration is produced with the sodium bicarbonate. If, for 
example, 19.8 cc. is the agreement, the solution would require 
the addition 0.2 cc. water to 19.8 cc. in order that 20 cc. corre- 
sponds to 10 cc. of the urea solution, but as the volume of the 



PREPARATION OF URINE FOR TITRATION. 123 

solution of sodium carbonate diluted the mixture titrated to a 
greater volume than 30 cc., a correction is necessary, which is 
made according to Pfliiger, by adding the number of cc. of the 
urea solution and sodium carbonate solution, and from the sum 
subtracting the number of cc. mercury solution required, and 
multiplying the remainder by 0.08, the product of which is sub- 
tracted from the number of cc. mercury solution employed in 
the titration. In this example, 13.85 cc. solution of sodium car- 
bonate was required to neutralize the acid — 

10 -f 13.85 = 23.85. 

Urea Sol. Na 2 C0 3 Sol. 

23.85 — 19.8 = 4.05. 4.05 x 0.08 = O.324. 19.8 — 0.324 = 19.476. 

It is seen from this correction that 19.476 cc. of the solution of 
mercuric nitrate would be required if the volume titrated did not 
exceed 30 cc. By the addition of 0.52 cc. water to 19.48 cc. the 
strength of the solution would be correct. To dilute 1 litre 26.6 
cc. water is required (19.48 : 0.52 : : 1000 : 26.69). The solu- 
tion will undergo no change in well-stoppered bottles kept in 
a dark place. 



PREPARATION OF THE URINE FOR TITRATION. 

Urine is prepared for titration by separating the phosphoric 
acid and chlorine. The former is separated first by means of the 
barium mixture, when the chlorine is separated from the filtrate 
by adding a definite quantity of the silver solution. To deter- 
mine the quantity of the silver solution required to separate the 
chlorine in 10 cc. urine, in which urea is to be estimated, titra- 
tions are made, employing Volhard's method, Chapter xi. This 
having been accomplished, 60 cc. of the urine is introduced into 
a dry 100 cc. flask from a burette, after which 20 cc. barium 
mixture is added. Mix well by shaking, and filter through a 
dry filter paper into a dry beaker or flask. Of the filtrate three- 
fourths is urine. For titrations, 60 cc. of the filtrate is introduced 
into a dry flask, and the number of cc. silver nitrate solution 
required to separate the chlorine in 45 cc. urine is added, as there 
is this quantity of urine in the 60 cc. Having mixed well by 
shaking, the solution is filtered through a dry filter paper into 
a dry flask. 



124 CHEMICAL ANALYSIS OF THE URINE. 

THE TITRATION. 

20 cc. of the filtrate from silver chloride is introduced into a 
small beaker and titrated as above in standardizing the mercuric 
nitrate solution, but as the quantity of urea in urine is subject to 
considerable variation, the result of the first titration is approxi- 
mative, after which within 0.2 or 0.3 cc. of the required quantity 
of the mercury solution is added without interruption, when the 
mixture is rendered nearly neutral with the soda solution, and 
the titrations continued as above. 

When the mixture is titrated with the solution of sodium car- 
bonate to neutralize the acid, blue litmus paper is employed to 
determine when a sufficient quantity has been added ; but as car- 
bonic acid is liberated in the reaction, and is partly held in solu- 
tion, it imparts the acid reaction after all of the nitric acid has 
combined with the sodium carbonate, for this reason, when the 
acid reaction of the mixture is slight, the indication is, that 
enough of the soda solution has been added. 

CALCULATION. 

If, at the time the titrations are completed (when the formation 
of the yellow coloration with sodium bicarbonate takes place), the 
volume of the fluid is more than 30 cc, correction is made as 
above, in standardizing the mercury solution ; in this case the 
sum of the volumes of urea solution (filtrate) and soda solution 
exceeds that of the mercury solution. If, on the other hand, the 
volume of the mercury solution is greater than that of the sum of 
the urea and soda solutions, the latter is subtracted from the 
former, the remainder multiplied by 0.08, and the product added 
to the number of cc. mercury solution. Having made the cor- 
rection, the volume of urine in the 20 cc. (of the filtrate) titrated 
is determined. Before the separation of chlorine from the first fil- 
trate three-fourths of its volume was urine, but the volume of silver 
solution required differs according to the quantity of chlorine pre- 
sent; hence, the number of cc. urine in 20 cc. of the filtrate (quan- 
tity titrated) is subject to variation. If, for example, the 45 cc. 
urine in 60 cc. of the filtrate required 37.4 cc. of the silver nitrate 
solution to separate the chlorine, then 20 cc. of the filtrate would 
contain 9.2 cc. urine (60 + 37.4 = 97.4 and ■— X 20 = 9.2). 



KNOP'S METHOD. 125 

As I cc. of the mercury solution (correction having been made) 
corresponds to o.oio grm. urea, the number of cc. mercury solu- 
tion employed multiplied by this number would give the number 

t t .1 • number grm. 

of grms. urea in 9.2 cc. urine, and by the proportion 9.2 : urea in 9 . 2 cc. 

urine 

: : 100 : x, the per cent, of urea in the urine is determined. 



ESTIMATION OF UREA IN DISEASED URINE. 

Albumen in urine interferes with the employment of the mer- 
curic nitrate solution in the same way that phosphoric acid does. 
It is removed by rendering a measured quantity of the urine — 
500 cc. — distinctly acid with acetic acid, boiling a few minutes 
in an evaporating dish, and to recover the loss by evaporation 
transfer to a T / 2 litre flask ; rinse the dish with a small quantity 
of water and add the rinsings to the urine in the flask. When 
cold, fill the flask with water to the mark, and having mixed well 
by shaking, filter through a dry filter paper into a dry flask, and 
separate the phosphoric acid and chlorine as if the urine were 
normal. Leucin and tyrosin combine with mercuric nitrate, but 
ordinarily the quantity of these bodies is so small that the results 
would differ but little by their absence. In cases of yellow 
atrophy of the liver, the quantity of leucin and tyrosin in the urine 
is so increased that this method cannot be employed, as there is 
no practical method for separating them from the urea. Sugar 
in the urine does not preclude the employment of this method. 

KNOP'S METHOD, MODIFIED BY GREENE. 

When urea in solution comes in contact with sodium hypo- 
bromite dissolved in water with sodium hydrate, nitrogen is set 
free — 

CH 4 ON 2 -f 3NaBrO = C0 2 + 2H 2 + 3NaBr + 2N. 
The C0 2 is absorbed by the sodium hydrate. The solution is 
prepared by dissolving 22 grms. sodium hydrate in 250 cc. water, 
and when cold adding 5 cc. bromine to the alkaline solution. The 
bromine is measured in a small graduated glass cylinder, and 
when emptied into the sodium hydrate solution, it is rinsed 
with the alkaline fluid and rinsings added to the hypobromite 
solution. The solution of hypobromite of sodium is kept in glass- 
stoppered bottles in a cool, dark place. 



126 



CHEMICAL ANALYSIS OF THE URINE. 



Fig. 32. 



Of the many forms of apparatus for the determination of urea 
by this method, that of Greene is the least complicated and 
admits of determinations without consuming much time. It is 
represented by Fig. 32, and consists of a graduated tube (a) of 20 
or 30 cc. capacity, and the lower extremity of 
which is enlarged and provided with a side 
tube which passes upward at an acute angle. 
The enlarged portion is drawn out and passes 
through a cork fitted into the opening in the 
receiver (c). The part (b) is a special pipette, 
having its lower end bent and provided with a 
cork. The graduated tube and enlarged por- 
tion are first completely filled with the hypo- 
brcmite and bromine solution and placed in 
the cup (c), when three or four cc. urine are 
introduced into the solution through the side 
tube by means of the pipette (b). The open- 
ing or orifice of the pipette is so small that at 
least a minute is required for the amount of 
urine to pass into the alkaline solution. In the 
course of thirty minutes the reaction is com- 
plete and nitrogen has collected in the gradu- 
ated tube. Either the pipette is fixed in place 
and water added until it stands on an even 
plane with the fluid in the graduated tube, or, 
for more accurate results, the pipette is removed and the gradu- 
ated tube (a) is placed in a vessel filled with water, so that the 
water and fluid in the tube are on an even plane, when the appa- 
ratus is secured in place by a holder and allowed to remain until 
the temperature of the air, water and nitrogen become the same. 
At the time of reading, the volume of nitrogen, the point at which 
the latter and the fluid meet, is brought even with the water in the 
cylinder or vessel. Memoranda of the temperature and baromet- 
ric pressure at the time of reading are made. The volume of dry 
nitrogen at zero C. and normal pressure, 760 mm., is determined 
by the following formula : — 




V 



V. P-T 



760 (1 -j- 0.00366 t) 



KNOP'S METHOD. * 127 

V = Volume of nitrogen in cubic centimetres. 

V = Number of cubic centimetres nitrogen read. 
P = Barometer reading. 

T= Vapor tension in millimetres, 
t = Thermometer reading. 

Vapor tension at ordinary temperatures is here given in milli- 
metres : — 



io°C. 


9.126 


15° C. 


12.677 


20°C. 


I7-396 


ii '• 


9-751 


16 " 


I3-5I9 


21 " 


I8.505 


12 " 


IO.42I 


17 " 


14.409 


22 " 


19.675 


13 " 


II. ISO 


18 " 


I5-35I 


23 " 


2O.9O9 


14 " 


11.882 


19 " 


16.345 


24 " 


22.211 



In one gramme of pure urea there is 371.4 cc. dry nitrogen at 
normal pressure and temperature ; but it has been found by this 
method that 354.3 cc. nitrogen is liberated from one gramme 
urea. This number is therefore employed in calculating the 
amount of urea from the volume of nitrogen, as shown by the 
equation, 354.3 : 1 : : nu ^ t b ro r g en. cc ' • x, when calculation is made of 
the quantity of urea in 100 cc, or in the total quantity of urine 
formed in twenty-four hours. 



CHAPTER X. 

Determination of the Total Quantity of Nitrogen in Urine — Dumas' Method — Var 
rentrapp and Will's Method — Kjeldahl's Method— Remarks on the Estimation of 
the Quantity of Urea and Nitrogen in Urine — Uric Acid — Salkowski's Method, 
modified by E, Ludwig — Kreatinin — Neubauer's Method, modified by Salkowski 
— Oxalic Acid — Neubauer's Method, modified by Fiirbringer. 



DETERMINATION OF THE TOTAL QUANTITY OF NITROGEN IN 

URINE. 

Of several methods for estimating the quantity of nitrogen in 
urine, three are here given — Dumas', Varrentrapp and Will's, and 
Kjeldahl's. By the method of Dumas, nitrogen is collected and 
measured, and by those of Varrentrapp and Will, and Kjeldahl, 
ammonia is formed and the quantity estimated by volumetric or 
gravimetric methods. 

DUMAS' METHOD. 

This method is based on the fact that organic bodies contain- 
ing nitrogen, when heated with copper oxide, are oxidized with 
the liberation of nitrogen, and any oxygen compounds of nitro- 
gen that may be formed are decomposed by metallic copper 
heated to redness. The weight of the nitrogen liberated is 
determined from the volume it would occupy at zero C. and 760 
mm. barometric pressure. The nitrogen is collected in a 200 
cc. graduated glass tube or cylinder with ground margins. The 
combustion tube employed is 70 to 80 cm. long with one end 
sealed by heating in the flame of a blast lamp. About 15 cm. 
of the sealed end is filled with dry sodium bicarbonate,* followed 
by about 10 cm. with finely granulated copper oxide, when a 
porcelain boat, containing the mixture of the solids of the urine 
and finely granulated copper oxide, is introduced. Formerly, 
urine, 5 cc, having been acidified with dilute sulphuric acid, was 
evaporated at ioo° C. to near dryness, and when cold mixed in 
a porcelain boat with finely granulated copper oxide by employ- 
ing a piece of wire, when the mixture was covered with copper 

* Pure dry manganese carbonate may be employed instead of sodium bicarbonate. 
It has the advantage of not liberating water by heating. 

128 



DETERMINATION OF NITROGEN IN URlNE. 129 

oxide and the boat introduced into the tube ; but the process has 
been shortened (Horbaczewski) by introducing 5 cc. urine into 
the porcelain boat half full of pulverized CuO, after which the 
boat is nearly filled with dry granulated CuO, when it is intro- 
duced into the combustion tube. Following the boat, finely 
granulated copper oxide is introduced, about 10 cm., when a roll 
of copper foil, 15 cm., is placed in the tube, and, finally, some 
more granulated CuO is introduced. The empty space of 5 cm. 
from the CuO to the end of the tube is cleansed with some por- 
ous paper, and by means of a tightly-fitting cork or rubber 
stopper connection is made with the delivery tube. To pre- 
vent the return of the water that may be formed in the open 
space at the end of the tube, the furnace is inclined somewhat. 
The graduated cylinder is filled two-thirds with mercury, air 
bubbles that may adhere to the glass are removed with a 
stirring rod, when it is filled to the margin with a strong 
solution of potassium hydrate. Slide a piece of ground glass 
over the ground surfaces of the cylinder, and if air bubbles are 
excluded invert the cylinder and place its open end, secured by 
the piece of ground glass, under the surface of mercury of the 
bath and retain the cylinder in place by a holder. About one- 
half of the sodium bicarbonate or manganese carbonate (6 cm. 
of the sealed end of the tube) is heated to redness. By decom- 
position of the NaHCO 3 carbonic acid is evolved, which drives 
the air out of the tube. 

2NaHC0 3 = H 2 + Na 2 C0 3 + CO,. 

To determine when this has taken place, collect some of the gas 
in a test tube filled with a solution of potassium hydrate over 
mercury from the delivery tube, and if the air has been driven 
out of the combustion and delivery tubes, and no air has been 
admitted from without, the gas will be completely absorbed. 
When this has taken place, the end of the delivery tube is placed 
under the mouth of the graduated cylinder, and the roll of copper 
foil in the combustion tube and the CuO adjacent thereto are 
heated to redness, and- the application of heat is continued 
toward the boat. The lamps of the furnace under the boat are 
not lighted until the water of the urine is evaporated by the heat 
of the CuO a few centimetres from the boat. This having taken 
9 



130 



CHEMICAL ANALYSIS OF THE URINE. 



place, apply heat gradually to the part of the tube in which the 
boat is until heated to redness. When complete combustion has 
taken place, known by no more nitrogen entering the graduated 
cylinder, the remainder of the sodium bicarbonate (or manganese 
carbonate) is heated gradually, that carbonic acid gas evolved 
drives out all of the nitrogen from the combustion and delivery 
tubes. The process having been completed and the delivery 
tube removed, the gas is turned off from the lamps, and the 
graduated tube having remained in position about thirty minutes, 
is transferred to a glass vessel, a large beaker glass for 
example, filled with water. This is brought about by placing a 
small evaporating dish under the mouth of the tube in the mer- 
cury, and while it is retained in the mercury in the evaporating 
dish it is removed to the vessel containing water. By removing 
the dish under the surface of the water, the mercury will sink to 
the bottom of the vessel and water will take its place. When 
the nitrogen, air and water are of the same temperature, the sur- 
face of the fluid in the cylinder is brought even with that of the 
surrounding water by lowering or raising the tube in the holder 
when the volume of nitrogen is read ; at the same time readings of 
the thermometer and barometer are made. Instead of employ- 
ing a simple graduated cylinder for collecting and measuring the 
nitrogen, Zulkowsky's azotometer (Fig. .33) may be employed. 
This apparatus comprises two tubes, A and 
B, each of which is 58 cm. long and 18 mm. 
diameter. Tube A is graduated, its upper 
end sealed, and by means of a piece of 
glass tubing welded in tube A, a few cen- 
timetres from the lower extremity, the rub- 
ber tube c and the small U-shaped tube 
d, it is connected with the combustion 
tube. Both tubes are filled with a strong 
solution of potassium hydrate, by first fill- 
ing the tube B, and then by inclining the 
apparatus the solution will run into tube 
A and the air pass out. This process is 
repeated until both tubes are filled, when some of the solution 
is allowed to escape through the tube secured by the pinch-cock 
b. If, when the first portion of the sodium bicarbonate is being 




VARRENTRAPP AND WILL'S METHOD. 131 

decomposed and the air driven out of the combustion tube, con- 
nection having been made with tube A, it is found that all of the 
gas which enters the tube is absorbed, the air has been expelled. 
If, however, air collects in the tube, it is removed by inclining 
the apparatus as in filling with the solution of sodium hydrate. 
During the combustion the pinch-cock a is removed, and that 
some unabsorbed carbonic acid may not cause an overflow of 
the tube B, some of the fluid is drawn off through the tube 
secured by the pinch-cock b. When the combustion has been 
completed, and all of the nitrogen driven out of the combustion 
and connecting tubes into tube A, the apparatus is disconnected, 
the tube again secured by the pinch-cock, and a strong solu- 
tion of potassium hydrate is introduced into tube B, and, after 
standing some time, a thermometer is introduced into the solu- 
tion in this tube, to ascertain the temperature. The pinch-cock 
b is opened to let sufficient sodium hydrate solution escape, to 
bring the fluid in both tubes even when reading of the barometer 
is made. 

CALCULATION. 

For the formula for the reduction of the volume of nitrogen, 
read to that of zero and 760 mm. pressure, refer to calculation of 
the quantity of urea from nitrogen, Knop and Greene's method, 
page 125 ; and having determined the volume of nitrogen at nor- 
mal temperature and pressure, multiply the volume (number ofcc.) 
by 0.001256, the product of which is the number of grammes 
nitrogen in 5 cc. urine. 

VARRENTRAPP AND WILL'S METHOD. 

When nitrogenous compounds of the urine are mixed with 
soda-lime (a compound of sodium and calcium hydrates) and 
heated to redness, the nitrogen combines with hydrogen of the 
hydrates, forming ammonia, NH 3 . The ammonia gas evolved is 
conducted through dilute sulphuric acid (normal sulphuric acid), 
by which it .is absorbed, forming (NH 4 ) 2 S0 4 . The combustion 
having been completed, the acid remaining uncombined is 
determined by titrating with a solution of potassium hydrate of 
known strength. For the preparation of normal sulphuric acid, 
refer to page 112, and to prepare normal potassium hydrate, 
page 109. The one-fifth normal potassium hydrate employed in 



132 CHEMICAL ANALYSIS OF THE URINE. 

titrating is prepared from the normal solution by introducing 
ioo cc. of the latter into a 500 cc. graduated flask and filling with 
distilled water to the mark. Soda-lime may be procured of deal- 
ers in chemical reagents. Unless it is free of nitrates or nitrites, 
it will be a source of error. To test for these impurities, mix the 
soda-lime with some pure cane sugar and heat in a tube sealed 
at one end to redness; ammonia is liberated, known by changing 
moistened turmeric paper dark red if nitrogen compounds are 
present. Soda-lime is dried before using by warming in a porce- 
lain dish. 

The combustion tube employed is about 40 cm. long and 12 
mm. in diameter, one end of which is drawn out to a fine point, 
sealed, and where the contraction begins it is bent so that the 
sealed extremity is several centimetres above the furnace. The 
sharp edges of the open end of the tube are rounded by heating in 
the flame of a blast lamp, care being taken not to heat sufficient to 
cause any irregularity in the opening. The absorption apparatus 
(Varrentrapp and Will's) comprises three bulbs communicating 
with each other by tubular openings. One of the bulbs is pear- 
shaped and is connected with the combustion tube by a glass 
tube passing through a cork fitting in the former. In filling the 
combustion tube, a tuft of asbestos is placed in the tube where the 
contraction begins, in front of which a small quantity of soda- 
lime — 4 to 5 cm. — is placed, when a mixture of the solids of the 
urine and soda-lime is introduced. For this purpose, 5 cc. urine 
and an equal volume saturated solution of oxalic acid are mixed 
in a small evaporating dish with a sufficient quantity powdered 
calcium sulphate (gypsum) to form a pasty mass, when it is dried 
at ioo° C. Instead of a dish a Hofmeister's capsule may be 
employed with advantage. This capsule is of very thin glass, so 
that it may be pulverized with its contents. In either case the 
dried mass is intimately mixed with some soda-lime in a mortar 
(avoiding heavy pressure with the pestle) and introduced into 
the combustion tube. The part of the mixture adhering to the 
mortar is removed by triturating small quantities of soda-lime, 
and adding to the mixture in the combustion tube. The part of 
the tube occupied by the mixture should be about 18 cm. in length. 
Finally, the tube is filled to 5 cm. of the end with soda-lime, and 
any particles of soda-lime in the open space are removed with 



VARRENTRAPP AND WILL'S METHOD. 133 

some soft paper, when a tuft of clean asbestos is fitted in the open 
space. By gently tapping the upper part of the tube space is 
formed for the passage of gases or vapors evolved. Into a 200 
cc. beaker introduce. 10 cc. normal sulphuric acid from a burette, 
and having placed the end of the tube of Varrentrapp and Will's 
absorption apparatus, connected with the round-like bulb, draw 
the acid into the bulbs by suction. The acid not being in suffi- 
cient quantity to fill the third or pear-shaped bulb half full when 
the apparatus is placed in the horizontal position, some water is 
introduced into the beaker with the acid yet remaining, and it is 
drawn into the bulbs as before. The beaker is then covered with 
a glass plate or large watch glass and placed one side, as it con- 
tains some of the 10 cc. normal acid. By means of a well-fitting 
rubber or cork stopper the absorption apparatus is connected with 
the combustion tube. To determine if the apparatus is air tight, 
warm the air in the pear-shaped bulb by holding a glowing coal 
or piece of hot iron near the surface of the glass, and by the 
expansion of the heated air a few bubbles will pass through the 
acid, and if the apparatus is air tight the acid in the pear-shaped 
bulb will remain at a higher plane than before. The anterior 
part of the tube containing the soda-lime is heated to redness, 
but care is taken not to heat the mixture until the soda-lime in 
front has reached a red heat ; at the same time gas in one or two 
lamps at the bend of the tube is turned on and ignited, so that the 
soda-lime contiguous to the tuft of asbestos be heated to redness 
and the drawn-out portion of the tube reach a degree of temper- 
ature sufficient to prevent the condensation of aqueous vapor. 
The application of heat is now continued slowly toward the 
sealed end of the tube until the tube is heated to redness from the 
bend to the tuft of asbestos in its anterior part. At no time dur- 
ing the process should the temperature of the soda-lime in the 
anterior part of the tube be reduced, neither should the heat be 
so intense as to incur the risk of decomposing the ammonia. The 
combustion is complete if, while the contents of the tube are heated 
to redness, the fluid in the bulbs is drawn toward the furnace. 
When this takes place,' the gas is turned off and air is drawn 
through the apparatus to cause the absorption of the ammonia 
remaining in the tube. This is brought about by connecting an 
aspirator with the free extremity of the bulbs, and the sealed end 



134 CHEMICAL ANALYSIS OF THE URINE. 

of the tube is broken by means of a pair of tongs. When about 
one litre water in the aspirator has been displaced by air (requiring 
about twenty-five minutes), the process is complete, and the 
quantity of acid in the bulbs is determined by titrating with the 
one-fifth normal potassium hydrate. For this purpose, the fluid 
is emptied into the beaker, which yet contains some of the 10 cc. 
normal acid, and after rinsing the bulbs several times with distilled 
water and adding the rinsings to the acid in the beaker, a sufficient 
quantity of solution of litmus (refer to page 114) is added to 
impart a distinct red color to the solution, when it is titrated with 
the one-fifth normal potassium hydrate from a burette until, by 
the addition of 0.1 cc, the color changes purple or blue. 

CALCULATION. 

In 1 cc. normal ammonia there is 0.017 g rm - NH 3 or 0.014 grm. 
nitrogen. 1 cc. normal acid will neutralize 1 cc. normal ammo- 
nia ; therefore, by multiplying the number of cc. normal acid 
neutralized by 0.017, the product is the quantity of NH 3 in 
grammes, or, by multiplying by p.014, the number of grammes 
nitrogen is determined. 

Example : 31 cc. of one-fifth normal potassium hydrate was 
required to neutralize the acid instead of 50 cc, as would be the 
case in the absence of ammonia. 3 1 cc. one-fifth normal solution 
is equal to 6.2 cc. of the normal p = 6.2), and as 10 cc. of the 
normal sulphuric acid was employed, 3.8 cc. was neutralized by 
the ammonia formed from 5 cc. urine (10 — 6.2 = 3.8), and. as 
3.8 cc normal ammonia would neutralize 3.8 cc. normal sulphuric 
acid, there is in 3.8 cc. normal ammonia the quantity of nitrogen 
found in 5 cc. urine, which is 0.0532 grm. (3.8 X 0.014 = 0.0532), 
and in 100 cc. urine there is 1.064 g rm - nitrogen (0.0532 X 20 = 
1 .064). 

KJELDAHL'S METHOD. 

This method is based on the fact that an organic body con- 
taining nitrogen, when heated with either fuming sulphuric acid 
or a mixture of fuming sulphuric acid and concentrated sul- 
phuric acid, sp. gr. 1.84, the nitrogen passes into the compound 
ammonium sulphate (NH 4 ) 2 S0 4 . By distilling a solution of the 
compound with an excess of sodium hydrate, ammonia is liber- 
ated, and in the receiver it is combined with a definite quantity 



kjeldahl's method. 135 

of sulphuric acid. When the process is complete, the amount 
of ammonia formed is determined by titrating the acid with 
one-fifth normal potassium hydrate. The method here given is 
nearly the same as that of Kjeldahl, as it is found accurate in 
estimating nitrogen in urine, blood and faeces ; but for the deter- 
mination of nitrogen in some other bodies it has been modified. 

SOLUTIONS REQUIRED. 

1. Fuming sulphuric acid. 

2. Normal sulphuric acid. 

3. One-fifth normal potassium hydrate. 

4. A solution of sodium hydrate, specific gravity 1.3. 

5. A solution of litmus. 

For the preparation of normal sulphuric acid, refer to page 
1 12. 

The one-fifth normal potassium hydrate is prepared by intro- 
ducing 100 cc. of the normal solution (for the preparation of which 
refer to page 109) into a 500 cc. graduated flask and filling with 
distilled water to the mark. To prepare a solution of sodium 
hydrate, sp. gr. 1.3, dissolve 320 grms. sodium hydrate in about 
500 cc. water, and, when cold, the. solution is introduced into a 
1000 cc. graduated flask and filled with water to the mark. As 
a high temperature is produced by dissolving sodium hydrate in 
water, place the flask or bottle containing the water and hydrate 
in cold water, and mix well by shaking, to prevent the parts of 
the glass in contact with the sodium hydrate from becoming so 
hot as to endanger the vessel. For the preparation of a solution 
of litmus, refer to page 114. 

APPARATUS REQUIRED. 

A 200 cc. round-bottom Bohemian flask. 

A 750 cc. Erlenmeyer flat-bottom flask, a condenser, and a 
400 cc. flask as a receiver, a safety tube, besides a 50 cc. burette, 
etc. 

THE ESTIMATION. 

Introduce 5 cc. urine from a burette into a 200 cc. round- 
bottom flask, and add 10 cc. fuming sulphuric acid.* 

To prevent loss while the fluid is heated, introduce (according 

* The fuming acid is conveniently measured in a small graduated glass cylinder or 
a test tube graduated for 10 cc. by a mark made with a sharp file. 



136 CHEMICAL ANALYSIS OF THE URINE. 

to Arnold) into the mouth of the flask a test tube which fits 
loosely into its neck, having been enlarged in its upper third by 
heating in the flame of a blast lamp and blowing out. If the 
enlargement of the test tube is near or in its middle third, remove 
the upper part of the tube with a file. Place the flask on wire gauze 
secured at an angle of 45 °, and heat the fluid with a gas or spirit 
lamp. Continue the application of heat until the fluid becomes 
light yellow in color. This usually takes place in one to one and 
one-half hours. The fluid should be kept in constant ebullition. 
After cooling, place the flask in cold water, as heat is generated 
by diluting, and add water in small quantities, mixing well by 
shaking gently after each addition, and when the dilution has 
reached about 100 cc. the fluid is introduced into the 750 cc. Erlen- 
meyer flask. Rinse several times with water, and add the rins- 
ings to the fluid. The quantity of fluid in the Erlenmeyer flask 
should not exceed 200 cc. Into the 400 cc. flask to receive the 
distillate, introduce 10 cc. normal sulphuric acid from a burette. 
The condensing tube of the cooler should be somewhat long, 
and the end to enter the receiver bent, that its orifice may be 
brought as near the acid as possible without coming in contact. 
As the fluid is dense when the solution of sodium hydrate is 
introduced, it is liable to bump during the distillation, to prevent 
which small fragments of zinc are introduced into the flask. By 
the action of NaOH on zinc, hydrogen is evolved, which prevents 
the bumping; but it was found (Pfeiffer and Lehmann) that with 
the hydrogen and aqueous vapor a small quantity of sodium 
hydrate is carried over, hence a safety tube is introduced between 
the Erlenmeyer flask and condenser. This is made by drawing 
out the end of a combustion tube, 20 cm. long and 18 mm. 
internal diameter, in the flame of a blast lamp to 8 or 10 mm. 
which passes through a hole in the cork of the Erlenmeyer flask.* 
The upper end of the tube is connected with the cooler by means 
of a cork, through which passes a bent glass tube. To the fluid 
to be distilled add 60 cc. of the solution of sodium hydrate (sp. 
gr. 1.30) and three fragments of zinc as nearly spherical as possi- 
ble, the weight of which not to exceed 0.5 grm. The sodium 

* x ~ It was found in this laboratory, that by a safety tube of the dimensions here 
given, the purpose is accomplished as well as with others more complex in construc- 
tion which have been recommended. 



ESTIMATION OF UREA AND NITROGEN. 137 

hydrate and zinc having been introduced, connection with the 
condenser is made at once to prevent loss of ammonia. Distill 
slowly until the ammonia separates from the fluid and is carried 
into the receiver and absorbed by the sulphuric acid. This is 
usually accomplished by distilling thirty minutes. To determine 
with greater accuracy if all the ammonia is distilled, place a 
small piece of red litmus paper at the orifice of the condensing 
tube, and if ammonia is still passing over, the red litmus paper 
turns blue. The quantity of ammonia absorbed by the 10 cc. 
normal sulphuric acid is determined by titrating with the one- 
fifth normal potassium hydrate. For this purpose, the solution 
of litmus is added to the fluid in quantity sufficient to impart a 
distinct red color, when the solution is titrated with the one-fifth 
normal potassium hydrate from a 50 cc. burette, until by the 
addition of 0.1 cc, after shaking, the solution turns purple or blue. 

CALCULATION. 

For the determination of the quantity of nitrogen from the 
quantity of acid neutralized in the titration, refer to calculation 
employed in Varrentrapp and Will's method, page 131. 

REMARKS ON THE ESTIMATION OF UREA AND NITROGEN. 

There is no method by which the exact amount of urea in 
urine is estimated, although the variation in results with some of 
the methods is not great. The difficulty is, other bodies in the 
urine either combine with, or are decomposed by, the reagent 
employed, according to the method. There is perhaps no method 
in quantitative analysis which has been more closely studied, and 
been the subject of discussion to such an extent, as Liebig's 
method, as variously modified, and although complicated, yet for 
clinical purposes it is satisfactory. To determine the amount of 
waste of the tissues, as in fever, exhaustive labor, or the waste 
from the transformation of nitrogenous elements of food in the 
blood, the most reliable methods are those by which all of the 
nitrogen in the urine is determined. For this purpose, the only 
objection, so far as the results are concerned, to Dumas' method 
is *the great difficulty, if not impossibility, of separating all of 
the air from copper oxide (Kreusler), and in very few cases, for 
example, the presence of nitro-substitution compounds in the 



138 CHEMICAL ANALYSIS OF THE URINE. 

urine, is there any objection to the method of Varrentrapp and 
Will. The results of Kjeldahl's method obtained in this labora- 
tory are entirely satisfactory, besides, the time saved by employ- 
ing this method is very great. Having the solutions prepared 
and the apparatus at hand, a half dozen estimations may be made 
during the time required to complete one estimation with the 
furnace. 

URIC ACID. 
SALKOWSKI'S METHOD, MODIFIED BY E. LUDWIG. 

This method of determining the quantity of uric acid in urine 
is based on the facts that silver urate is insoluble in solutions 
rendered strongly alkaline with ammon. hydrate, and that when 
suspended in water it is decomposed with sodium sulphide, form- 
ing silver sulphide and sodium urate. The phosphates of cal- 
cium and magnesium are separated with the silver sulphide by 
filtering. 

For the employment of this method, three solutions are 
required: A solution of silver .nitrate, an alkaline solution — 
ammonia solution — of the double chloride of magnesium and 
ammonium or " magnesia mixture," and a solution of sodium 
sulphide. 

Dissolve 26 grms. crystallized silver nitrate in 200 or 300 cc. 
distilled water in a 1000 cc. graduated flask, add an excess of 
ammon. hydrate and with water fill to the mark. Mix well by 
shaking. 

For the preparation of magnesia mixture, refer to page 115. 

Dissolve 10 grms. sodium hydrate in 1000 cc. water. Mix well 
by shaking, and treat 500 cc. of the solution with sulphureted 
hydrogen gas to saturation, when the two solutions are mixed, 
forming sodium sulphide. 

NaOH + H 2 S = NaHS + H 2 and NaHS + NaOH = Na 2 S + H 2 0. 

Having prepared the three solutions, introduce 25 cc. of the 
silver solution and an equal volume of the magnesia mixture into 
a flask, add ammon. hydrate until the precipitate AgCl dissolves. 
This mixture is added to 200 cc. filtered urine and mixed well 
with a glass rod. After the lapse of an hour filter through a filter 
secured by a platinum cone, and to facilitate the process connec- 
tion is made with a filter pump or an aspirator bottle. Wash 



KREATININ. 139 

the precipitate three times with water containing some ammon. 
hydrate. Transfer the precipitate to the beaker in which the pre- 
cipitation took place by means of a stream of water from a wash 
bottle, taking care, however, not to injure the filter. To the pre- 
cipitate, suspended in the water, the sodium sulphide solution is 
added in sufficient quantity to precipitate the silver as sulphide. 
when the mixture is heated until the boiling point is reached, 
while it is constantly stirred. After cooling, filtert hrough the 
filter employed before into an evaporating dish, and wash with 
hot water until a sodium hydrate solution of lead acetate ceases 
to produce a precipitate or dark coloration with the wash water. 
Render the filtrate with wash water in the dish faintly yet dis- 
tinctly acid with dilute hydrochloric acid, and concentrate by 
evaporating on a water bath to the volume of about 1 5 cc. Let 
stand three hours in a cool place, when the uric acid will separate. 
Filter through a small, weighed filter paper, having been dried at 
no° C. The filtrate is employed in connection with a stirring 
rod, over the end of which is placed a short piece of rubber tube, 
in bringing all of the uric acid on the paper, when it is washed 
several times with water and dried in the funnel, but at a temper- 
ature not exceeding 1 io° C. The funnel and filter having cooled, 
the uric acid is washed with about 10 cc. carbon bisulphide in 
three portions, after which wash with a small quantity of ether 
and dry at no°, and after cooling in a desiccator, weigh. The 
process of heating, cooling and weighing is repeated until the 
weight becomes constant. The difference between the weight of 
the filter and that of the filter + precipitate is the weight of uric 
acid in 200 cc. urine, or twice the per cent, of uric acid in the urine, 
not taking the specific gravity of the urine into consideration. 

KREATININ. 
NEUBAUER'S METHOD, MODIFIED BY SALKOWSKI. 

After the separation of phosphoric acid the urine is evaporated 
to dryness, an alcohol solution of constituents of the residue 
soluble in alcohol prepared, and the kreatinin precipitated with an 
alcohol solution of zinc Chloride. To prepare the alcohol solution 
of zinc chloride, the syrupy chloride is treated with strong alcohol 
until the specific gravity of the solution becomes 1.2, when it is 
filtered. Introduce 240 cc. of the urine into a graduated 300 cc. 



140 CHEMICAL ANALYSIS OF THE URINE. 

flask, render slightly alkaline with milk of lime, and treat with a 
solution of calcium chloride until a precipitate ceases to form. 
Fill the flask with water to the mark and mix well by shaking. 
After standing fifteen minutes, filter through a dry filter paper. 
The filtrate should be alkaline in reaction ; but if decidedly so, 
neutralize with hydrochloric acid, but not until 250 cc. of the filtrate 
is introduced into an evaporating dish. Evaporate on a water bath 
to about 20 cc, and when cold add an equal volume of absolute 
alcohol. Mix well by stirring with a glass rod, and introduce the 
fluid into a graduated 100 cc. flask. Rinse the dish several times 
with absolute alcohol and add the rinsings to the fluid in the 
flask, and fill the flask with absolute alcohol to the mark. Mix 
well by shaking after having fitted the stopper in place. Having 
stood twenty-four hours, filter through a dry filter paper and 
introduce 80 cc. of the filtrate into a beaker glass. To the alco- 
hol solution add about 1 cc. of the alcohol solution of zinc chlo- 
ride ; mix well by stirring with a glass rod several minutes ; finally 
cover with a glass plate or large watch glass and let stand in a 
cool place two or three days. The solution is then filtered through 
a small, weighed filter paper which was dried at ioo° C. The 
filtrate is employed in collecting and bringing the precipitate on 
the filter by means of a stirring rod, the end of which is provided 
with a small piece of rubber tubing. Wash the precipitate with 
small quantities of strong alcohol until the alcohol filtrate is free 
of chlorine, known by testing with a solution of silver nitrate 
and a few drops of dilute nitric acid. When washed, the filter 
paper with precipitate is dried and weighed as before. 

CALCULATION. 

The difference in weight between the filter with watch glasses 
(between which the filter was weighed) and the filter -j- precipitate 
with watch glasses, is the weight of kreatinin zinc chloride. 
240 cc. urine is diluted to 300 cc, and when filtered, 250 cc. 
of the filtrate is taken, which corresponds to 200 cc. urine 
(^g X 240 = 200). Of the 100 cc. alcohol solution, 80 cc is 
taken, and as the 100 cc solution contains the kreatinin of 200 
cc urine, 80 cc. contains the kreatinin of eight-tenths as much 
urine, which is 160 cc, and five-eighths of this quantity of krea- 
tinin corresponds to 100 cc urine (^- = $4); therefore, by mul- 



OXALIC ACID. 141 

tiplying the weight of kreatinin zinc chloride, weighed, by five- 
eighths, the product of which is the weight of the compound from 
ioo cc. urine, and as 62.44 P er cent - °f tne zmc compound is 
kreatinin, by multiplying by 0.6244, tne product is the weight 
of kreatinin in 100 cc. urine. 



OXALIC ACID. 

NEUBAUER'S METHOD, MODIFIED BY FURBRINGER. 

The quantity of urine passed in twenty-four hours is measured 
and treated with a few cc. oil of thyme, to prevent bacterial 
growth. Treat the urine with ammon. hydrate until, after stirring, 
the odor of ammonia is perceptible. Add a solution of calcium 
chloride until a precipitate ceases to form, and with acetic acid 
render distinctly acid, avoiding a great excess. The phosphates 
of calcium and magnesium dissolve in acetic acid, while calcium 
oxalate precipitates with more or less uric acid. Let stand twen- 
ty-four hours, filter through a small filter paper, collect and trans- 
fer the precipitate to the paper with a glass rod provided with a 
small piece of rubber tubing on one end. Wash with water until 
the wash water is free of chlorides, known by testing with a solu- 
tion of silver nitrate and a few drops of nitric acid. When washed, 
transfefthe filter, with precipitate, to a small beaker and treat with 
dilute hydrochloric acid and water, avoiding a great excess of the 
former; warm on a water bath, and stir with a glass rod so the 
acid will come in contact with every part of the precipitate. The 
calcium oxalate will dissolve in the acid, while any uric acid pres- 
ent will remain undissolved. Filter, through a small filter paper, 
into a beaker of 250 or 300 cc. capacity, wash with water and 
determine when washed, as before. Evaporate the filtrate with 
wash water to about 200 cc, transfer from the dish to a beaker, 
rinse the dish with water and add rinsings to the fluid in the 
beaker when the fluid is rendered alkaline with ammon. hydrate, 
known by turning turmeric paper dark red after the fluid is well 
mixed by stirring. Having stood well protected from dust twen- 
ty-four hours, filter through a small filter paper free of ash, wash 
with water until the wash water is free of chlorine, known by 
producing no turbidity when tested with a solution of silver 
nitrate with a few drops of nitric acid. Dry the filter with pre- 



142 CHEMICAL ANALYSIS OF THE URINE. 

cipitate and ash in a platinum crucible, and by the heat of a good 
blast lamp reduce the oxalate to the oxide (CaO), let cool in a 
desiccator and weigh. Repeat the process of heating and weigh- 
ing until the weight becomes constant. To find the weight of the 
oxalic acid,- C 2 H 2 4 , multiply the weight of the oxide of calcium, 
CaO, by 1.607 1. 



CHAPTER XL 

Phosphoric Acid of the Phosphates and of Glycerin- Phosphoric Acid — Neubauer and 
Zuelzer's Method — Phosphoric Acid of the Phosphates — Total Quantity of Phos- 
phoric Acid — Sulphur Compounds — Sulphuric Acid or Sulphur of Sulphates and 
Ester Compounds — The Gravimetric Method — The Volumetric Method (Wilden- 
stein, Briiggelmann and Neubauer) — Sulphuric Acid of the Ester Compounds — 
Sulphur not in Sulphates or Ester Compounds — Method of Estimating Total Quan- 
tity of Sulphur in Urine. — Chlorine, Volhardt's Method, Modified by Salkovvski — 
Neubauer' s Method — Remarks on the Methods of Estimating Chlorine in Urine — 
Potassium and Sodium — Ammonia, Schlosing and Neubauer's Method — Calcium 
and Magnesium Oxides — Calcium Oxide — The Gravimetric Method — The Volu- 
metric Method (Neubauer) — Magnesium Oxide — The Gravimetric Method — The 
Volumetric Method (Neubauer). 



PHOSPHORIC ACID OF THE PHOSPHATES AND OF GLYCERIN- 
PHOSPHORIC ACID. 

NEUBAUER AND ZUELZER'S METHOD. 

Phosphorus in the urine is in the form of simple phosphates, 
and in an organic body glycerin-phosphoric acid (refer to page 
34). The latter body is decomposed by heating with dilute 
nitric acid, with the liberation of phosphoric acid, which is pre- 
cipitated by ammon. hydrate and magnesia mixture. Without 
this preliminary treatment the glycerin-phosphoric acid would 
remain in solution ; hence, by means of the magnesia mixture, the 
two phosphorus compounds are separated. The magnesium 
ammon. phosphate, MgNH 4 P0 4 , precipitated by the magnesia 
mixture, whether of the phosphates or of the total quantity of 
phosphorus in the urine, is filtered, washed and transferred to a 
beaker, dissolved in acetic acid and titrated with a solution of 
uranium acetate of known strength. Formerly, the urine was 
titrated directly with the uranium solution — free acetic acid pres- 
ent — but by the researches of Zuelzer the results were found too 
high. 

SOLUTIONS REQUIRED. 

1. Magnesium mixture. 

2. Solution of sodium phosphate. 

3. Solution of uranium acetate. 

4. Solution of sodium acetate. 

143 



144 CHEMICAL ANALYSIS OF THE URINE. 

PREPARATION OF SOLUTIONS. 
For the preparation of magnesium mixture, refer to page 115. 

SOLUTION OF SODIUM PHOSPHATE. 

From a solution of sodium phosphate of known strength a 
solution of uranium acetate is standardized. The solution for 
this purpose contains 10.0845 grms. sodium phosphate, Na 2 HP0 4 , 
I2H 2 0, in 1000 cc. water or 0.2 grm. P 2 5 in 100 cc water. The 
pure phosphate, with water of crystallization, may be weighed, put 
into a dry flask or bottle, and, by calculation, the required amount 
of water be added. For example, if 6.2831 grms. of the salt is 
weighed on a balance of precision, the quantity of water in which 
to dissolve the salt is calculated by the equation 10.0845 : 1000 
: : 6.2831 : x (x = 623.4 cc). With greater accuracy, about 
7 grms. pure sodium phosphate is dissolved in 600 cc. water, and 
when dissolved and well mixed by shaking, 50 cc. of the solution 
is evaporated to dryness in a weighed platinum dish, and with the 
cover on the dish it is gradually heated until the bottom of the 
dish becomes dull red in color. Having cooled in a desiccator, it 
is weighed ; the increase in weight is due to sodium pyrophos- 
phate, Na 4 P 2 7 , formed by the decomposition of the ordinary 
phosphate by heat. The quantity of the sodium pyrophosphate 
in 50 cc. of the solution corresponding to the quantity of sodium 
phosphate required is 0.1873 g rm -> an d 500 cc. of the solution is 
diluted accordingly. If, for example, 50 cc. of the solution yields 
0.1950 grm. of the pyrophosphate, the quantity of water with 
which to dilute 50 cc. of the solution is determined by the equa- 
tion 0.1873 : 50 : : 0.1950 : x. x = 52.05. If 50 cc. requires 
2.05 cc. water, 500 cc. would require 20.5 cc. (10 X 2.05 = 20.5). 

SOLUTION OF URANIUM ACETATE. 

About 44 grms. uranium acetate is dissolved in 1100 cc. water, 
or 35 grms. of the oxide of uranium is dissolved in acetic acid 
and the solution diluted with water to 1100 cc. This solution is 
standardized with the sodium phosphate solution, as directed 
below. 

SOLUTION OF SODIUM ACETATE. 

Dissolve 100 grms. pure sodium acetate in water, introduce the 
solution into a 1000 cc. graduated flask, and fill with water to the 
mark. 



PHOSPHORIC ACID OF THE PHOSPHATES. 145 

SOLUTION OF POTASSIUM FERROCYANIDE. 

io grms. of potassium ferrocyanide is dissolved in ioocc. water. 

STANDARDIZING THE SOLUTION OF URANIUM ACETATE. 

When a solution of uranium acetate is brought in contact with 
a solution of sodium phosphate a light yellow precipitate forms — 
2UOC 2 H 3 2 + Na 2 HP0 4 = (UO) 2 HP0 4 -f 2NaC 2 H 3 2 . 

If the uranium solution be added to the phosphate, and there be 
mixed with the latter some potassium ferrocyanide and sodium 
acetate in solution, uranium ferrocyanide (a dark-colored precipi- 
tate) will not form unless the phosphoric acid has combined with 
uranium, and there is in the solution an excess of uran. acetate. 
In titrating, therefore, potassium ferrocyanide is used as the indi- 
cator. In titrating solutions in which there is an acid beside 
acetic acid, the sodium acetate is added that the sodium of the 
acetate combine with the acid (or the acid of an acid salt, if pres- 
ent) and acetic acid be liberated. 

Into a flask of 150 or 200 cc. capacity introduce from a pipette 
50 cc. of the solution of sodium phosphate, add 5 cc. of the solu- 
tion of sodium acetate, heat to the boiling point and titrate the hot 
solution with the uranium solution from a burette until a precipi- 
tate ceases to form. Transfer a drop of the hot mixture by means 
of a stirring rod to a porcelain dish, in which there is a drop of 
the solution of potassium ferrocyanide, and when the two drops 
are brought together, if no dark coloration takes place, heat the 
mixture again,. titrate with the uranium solution and test as before. 
Repeat the process until by testing a dark color is produced. 
When an agreement is reached, the uranium solution is diluted 
so that 20 cc. corresponds to 50 cc. of the sodium phosphate 
solution. If, for example, 18.2 cc. of the uranium solution is 
sufficient, the addition of 1.8 cc. water would be required, and 50 
cc. would require 49.4 cc. water (18.2 : 1.8 : : 500 : x). 1 cc. of 
the standardized uranium solution corresponds to 0.005 g rm - P2O5. 

DETERMINATION OF THE PHOSPHORIC ACID OF THE PHOSPHATES. 

If the urine is not clear it is filtered, or, if the urine is alkaline, 
neutral, or slightly acid, and it is desired to include the phos- 
phoric acid of the sediment in the estimation, render the urine 



146 CHEMICAL ANALYSIS OF THE URINE. 

strongly acid with acetic acid; mix well by stirring with a glass 
rod and filter, if necessary. Introduce 500 cc. of the urine in a 
beaker, add 10 cc. solution ammon. chloride (1 part.ammon. chlo- 
ride and 10 parts water) and 40 to 50 cc. magnesium mixture, 
when 100 cc. strong ammon. hydrate is added and the fluid well 
mixed with a stirring rod. Having stood 6 to 12 hours, filter, 
and transfer the precipitate to the filter by means- of a stirring 
rod provided with a small piece of rubber tubing placed on its 
end. Wash the precipitate with water containing one-third its 
volume of ammon. hydrate. The washing is continued until 
some of the wash water, having been boiled in a test tube to drive 
off the excess of ammonia, and rendered acid with nitric acid, 
ceases to yield a turbidity with a solution of silver nitrate. When 
the precipitate is washed, it is immediately transferred to a gradu- 
ated 250 cc. flask. This is brought about by perforating the filter 
with a glass rod, and with a fine stream of water from a wash 
bottle every trace of the precipitate is removed from the paper. 
Dissolve the precipitate with acetic acid, and with water fill to the 
mark. Mix well by shaking. With a pipette introduce 50 cc. 
of the solution into a 200 cc, flask, add 5 cc. of the solution of 
sodium acetate, heat to the boiling point (preferably on a sand 
bath), and from a burette containing the uranium solution titrate 
while hot and test with the solution of potassium ferrocyanide as 
in standardizing the uranium solution. Titrations of other por- 
tions of 50 cc. are made until an agreement is reached. 

DETERMINATION OF THE TOTAL QUANTITY OF PHOSPHORIC 

ACID. 

The quantity of P 2 5 in glycerin-phosphoric acid, as well as 
that of the phosphates, is determined by boiling the urine, 
having been acidified with nitric acid. For this purpose 250 cc. 
urine is rendered strongly acid with nitric acid and boiled thirty 
minutes, and, when cold, the phosphoric acid is precipitated by 
rendering the solution alkaline with ammon. hydrate and treat- . 
ing with 50 cc. magnesium mixture and ammon. hydrate, when 
the estimation is continued as above in determining the quantity 
of P 2 5 of the phosphate. 



SULPHURIC ACID .AND ESTER COMPOUNDS. 147 

CALCULATION. 

20 cc. of the uranium solution corresponds to o.i grm. P 2 5 . 
If more or less than 20 cc. of the uranium solution were em- 
ployed for 50 cc. of the solution, 25 cc, for example, the equation 
would be 20 : 0.1 : : 25 : x. x = the weight of P 2 5 in 50 cc. 
of the solution, or multiply the number of cc. uranium solution 
employed in the titration by 0.005, the product of which is the 
weight of P 2 5 in 50 cc. of the solution. The number 0.005 i s 
the weight in grms. of P 2 5 , which corresponds to 1 cc. of the 
uranium solution. As the volume of the solution titrated cor- 
responds to the volume of urine, multiply the number of grms. 
P 2 5 found in 50 cc. by two, the product of which is the per 
cent, of P 2 5 in the urine, not taking into consideration the 
specific gravity of the urine. The difference between the weight 
of P 2 5 of the phosphates and the weight of the total quantity 
of P 2 5 in the urine is the weight of P 2 5 of the glycerin-phos- 
phoric acid. 

SULPHUR COMPOUNDS. 

For the combinations of sulphur in the urine, refer to page 36. 

The estimations of sulphur and sulphuric acid in the urine 
are(i) the quantity of sulphuric acid of both sulphates and ester 
compounds ; (2) the quantity of sulphuric acid of ester com- 
pounds, and (3) the quantity of sulphur of compounds other than 
sulphates and esters. 



SULPHURIC ACID OR SULPHUR OF SULPHATES AND ESTER COM- 
POUNDS. 

THE GRAVIMETRIC METHOD. 

The gravimetric method of determining the quantity of sul- 
phuric acid or sulphur in the urine differs in some respects from 
that employed in estimating it in aqueous solutions, as the barium 
sulphate forming in the urine is liable to incorporate bodies 
which cannot be separated by washing with water. It is, there- 
fore, recommended to dilute 50 cc. urine to 150 cc. with water 
before precipitating with barium chloride. If the urine is not 
clear, it is filtered through a dry filter into a dry flask before 
diluting, when 50 cc. is introduced into a 250 or 300 beaker 
with a pipette and diluted with 100 cc. water. x'Acidify with a 



148 CHEMICAL ANALYSIS OF THE URINE. 

small quantity of hydrochloric acid, heat on a wire gauze until the 
fluid begins to boil, and precipitate with a clear solution of 
barium chloride (i part barium chloride and 10 parts water). 
When precipitated, place the beaker on a water bath ; the precipi- 
tate having settled, determine if the precipitation is complete by 
introducing a small quantity of the solution of barium chloride into 
the warm fluid with a pipette ; if no cloudiness is produced, enough 
of the barium solution has been added. If, on the other hand, a 
precipitate forms, heat again on a wire gauze and add more of 
the solution of barium chloride. Complete precipitation having 
taken place, the mixture is heated one or two hours on a water 
bath, and filtered by decanting through a small, fine filter paper 
(preferably washed Swedish filter paper or No. 590 Schleicher 
and Schull's filter paper). When the fluid has been decanted, 
except 20 or 30 cc, add about 50 cc. distilled water, stir with a 
glass rod without touching the glass, heat again on the water 
bath until the precipitate has settled, and filter as before. Collect 
the precipitate on the filter, and remove every trace that may 
remain in the beaker with a stirring rod provided with a small 
piece of rubber tubing placed over its end. Wash with distilled 
water until the wash water ceases to yield a precipitate with a 
solution of silver nitrate, then continue the washing with about 
100 cc. alcohol heated in a flask to the boiling point. The alcohol 
will remove some of the organic matter insoluble in water. 
With the precipitate there are still impurities, to separate which 
(Briiggelmann) dry the filter with precipitate at ioo° C, and 
transfer the precipitate to a platinum crucible or small platinum 
dish and ash the paper on a platinum wire. The precipitate, 
with the ash, is moistened with concentrated hydrochloric acid, 
when 3 or 4 cc. water is added, and the mixture well stirred with 
a short stirring rod. Warm gently over the free flame and 
decant the fluid through a small filter. Repeat this process five 
times, then transfer the precipitate to the filter and wash with 
water until the wash water ceases to yield a precipitate or cloudi- 
ness with a solution of silver nitrate. Dry the precipitate with 
filter at ioo° C. and ash* the paper, and into a weighed platinum 
crucible introduce the precipitate and ash, and heat ten minutes 
with a Bunsen's burner or Argand spirit lamp. After cooling, 
moisten with dilute sulphuric acid and heat gradually to dull 



THE VOLUMETRIC METHOD. 149 

redness, place in a desiccator and, when cool, weigh. The treat- 
ment with dilute acid is, to transform any barium sulphide, that 
may have formed, into the sulphate. 

CALCULATION. 

The molecular weight of BaS0 4 is 233, that of H 2 S0 4 is 98 ; 
the atomic weight of sulphur is 32, therefore, 



Df : x. x = weight of H 2 SO, and 
233 - 3 2 '■ : Baso* : x. x =r weight of Sulphur. 



*JJ ■ *" •■ BaS0 4 
Wt. of 



The quantity of H2SO4 or sulphur found, is from 50 cc. urine, 
therefore, for 100 cc. multiply by two. 



THE VOLUMETRIC METHOD. 
WILDENSTEIN, BRUGGELMANN AND NEUBAUER. 

The solutions required for the volumetric method of estima- 
ting sulphuric acid are one-fifth normal solutions of barium chlo- 
ride and potassium sulphate, and the apparatus required is a 
special filter besides burettes. 

ONE-FIFTH NORMAL BARIUM CHLORIDE. 

Pure barium chloride, containing water of crystallization, is 
dried by pressing between filter paper, weighed on a balance of 
precision, put into a clean dry flask of suitable size, and water is 
added, so that in 1000 cc. of the solution there are 24.392 grms. 
barium chloride. The quantity of water in which to dissolve the 
salt is determined by the equation — 

24.392 : 1000 :; BaS'tweTJhed : *• * — No. cc. water required. 
ONE-FIFTH NORMAL POTASSIUM SULPHATE. 

To prepare the solution of potassium sulphate, the salt, chemi- 
cally pure, is dried at ioo° C, and, having cooled in a desiccator, 
it is weighed and introduced into a clean, dry flask. Add water 
to the salt in quantity until the strength of the solution is 17.4 
grms. potassium sulphate in 1000 cc. of the solution. From 
the weight of the saltlhe quantity of water in which to dissolve 
the salt is determined by the equation — 

17.4 : 1000 :: Kfs 6 4 °S'ed : x - x = No - cc - water required. 



150 



CHEMICAL ANALYSIS OF THE URINE. 



Fig. 34. 



The special or syphon filter required is represented by Fig. 
34. It is made of a piece of combus- 
tion tubing, the internal diameter of 
which is 12 to 15 mm., and drawn out 
from 15 to 20 mm. of one end by heat- 
ing in a blast lamp successive parts until 
the length of the drawn-out part is 17 
cm. long, then the tube is bent about 8.5 
cm. from the bell-shaped end, as seen by 
the Fig. A small rubber tube passes 
over the lower end of the syphon and 
extends to a lower plane than the end 
with the filter. The rubber tube is 
secured by a pinch-cock. The filter in 
the bell-shaped end of the tube is made 
of well-washed filter paper. One layer is placed over the con- 
stricted part, followed by fragments of wet paper. Before using, 
water is drawn through the filter to ascertain if the packing is 
loose enough to permit the passage of water, and if it is found 
that the water passes through with difficulty, less material is 
employed or the fragments of paper are loosened. 




THE TITRATION. 

100 cc. of the filtered urine is introduced into a beaker of 200 
to 300 cc. capacity and rendered acid with 2 cc. hydrochloric 
acid, sp. gr. 1.12. The beaker is then placed on a wire gauze or 
sand bath and heated to the boiling point, when portions of 1 cc. 
of the one-fifth normal barium chloride are added from a 25 cc. 
burette until, after mixing with a stirring rod, no further precipi- 
tation appears to form. To the end of the process the tempera- 
ture of the mixture is kept at the boiling point. The end of the 
syphon with the filter is introduced into the mixture, and while 
the other extremity is raised and turned, a glass tube is intro- 
duced into the rubber tube, and the urine is drawn by suction 
through the filter nearly to the end of the syphon, when the pinch- 
cock is attached and the filter is ready for use. Filter about 
2 cc. of the fluid into a test tube and add a few drops of the 
barium solution from the burette. If a precipitate forms, the fluid 



THE TITRATION. 151 

in the test tube is returned to the beaker, and the test tube is 
rinsed with water two to three times and the rinsings added to 
the fluid in the beaker. Add I cc. of the' barium solution, mix 
well with a stirring rod, filter 3 or 4 cc., but instead of testing 
this filtrate, return it to the beaker, as this quantity of fluid may 
have been in the tube during the last titration. Having mixed 
well, filter another portion and test with the barium solution ; 
if a precipitate or cloudiness is produced, repeat with 1 cc. of 
the barium solution until a precipitate is no longer produced. 
Having determined about how much of the barium solution is 
required for 50 or 100 cc. urine, empty and clean the beaker, 
wash the filter with distilled water and fill the burette to the o 
mark with the barium solution. Introduce the same quantity of 
urine and acid into the beaker, heat and titrate with 1 cc. less of 
the barium solution than was employed before. Stir, filter, and 
test the filtrate with 0.1 cc. of the barium solution. If no cloudi- 
ness is produced, titrate with 0.1 cc. of the barium solution, and 
continue the titrations, adding o.l cc. of the barium solution 
both for titrating and testing, until the filtrate, when tested, 
becomes cloudy after the lapse of a few seconds, when another 
portion is filtered and tested with the one-fifth normal potassium 
sulphate, and if a precipitate is produced immediately, too much 
of the barium chloride solution has been added, in which case 
another titration is made with the same quantity of urine, but 
employing 0.3 cc. less of the solution of barium chloride, and if, 
by testing, a light precipitate forms at once, titrate with 0.1 cc. of 
the barium solution until a light precipitate forms in the filtrate 
several seconds after the addition of the solution of barium chlor- 
ide. When this takes place, another portion is filtered and tested 
with the solution of potassium sulphate, and if the fluid becomes 
somewhat turbid in 15 to 20 seconds, the exact quantity of the 
solution of barium chloride required by the sulphuric acid in the 
urine has been employed. 

CALCULATION. 

Multiply the number of cc. of the one-fifth normal barium 
chloride employed by 0.0098; the product is the number of 
grms. H 2 S0 4 ; or, for the quantity of sulphur in the quantity of 
urine titrated, multiply the number of cc. of the one-fifth normal 
barium chloride employed by 0.0032. 



152 CHEMICAL ANALYSIS OF THE URINE. 

SULPHURIC ACID OF THE ESTER COMPOUNDS. 

For the estimation of sulphuric acid of the ester compounds 
in the urine, that of the sulphates is first separated. For this 
purpose saturated solutions of barium hydrate and barium chlo- 
ride are prepared, when two volumes of the former are mixed 
with one volume of the latter, ioo cc. of the urine and ioo cc. 
of the barium solution or mixture are introduced into a dry 
beaker and well mixed with a stirring rod, and in 10 or 15 
minutes the solution is filtered through a dry filter into a dry 
beaker. Introduce 100 cc. of the filtrate into a beaker, render 
strongly acid with hydrochloric acid and heat to the boiling 
point, then proceed in estimating the sulphuric acid by the 
gravimetric method, as above. 



SULPHUR NOT IN SULPHATES OR ESTER COMPOUNDS. 

To determine the quantity of sulphur in the urine, not in 
sulphates or ester compounds, estimations are made of the 
sulphur of the latter compounds, as above, by the gravimetric or 
volumetric method, and of the total quantity of sulphur in the 
urine, and from the weight of the latter the weight of the former 
is subtracted ; the remainder is the weight of the sulphur in com- 
pounds, the constitution of the greater number of which is 
unknown. 

To estimate the total quantity of sulphur in urine, introduce 
50 cc. of the urine into a platinum dish, render strongly alkaline 
with a solution of sodium carbonate and add 3.5 grms. potassium 
nitrate, when the dish is placed on a water bath and the evapora- 
tion continued until the water is driven off. Heat is then applied 
gradually until the residue fuses, avoiding any higher tempera- 
ture than necessary to bring about fusion. If the fusion is still 
dark in color, small crystals of potassium nitrate are added and 
the heat continued until it becomes white. When cold, treat the 
residue with water and separate the insoluble part BaC0 3 by 
filtering, wash with water and add wash water to the filtrate. The 
residue is washed until the wash water ceases to change the color 
of turmeric paper. To transform the nitrates and nitrites in the 
filtrate and wash water, evaporate in a porcelain dish on a water 
bath with an excess of hydrochloric acid. To drive off the nitric 



CHLORINE. . 153 

and nitrous acids, the process is repeated, with the addition of a 
small quantity of hydrochloric acid. Dissolve the residue in 
water, transfer the solution to a beaker, rinse the dish several times 
with water and add the rinsings to the fluid in the beaker. The 
sulphuric acid in the fluid may be estimated either by the volu- 
metric or gravimetric method ; if by the latter, the process may 
be shortened somewhat, as it is not necessary to wash the BaS0 4 
with alcohol or treat it with dilute sulphuric acid. 

CALCULATION. 

For the calculation of the quantity of sulphur from the weight 
of BaS0 4 , refer to calculation of the gravimetric method. If the 
volumetric method has been employed, for the quantity of sul- 
phur, multiply the weight of H 2 S0 4 by - % . The weight of the 
sulphur of the sulphates and ester compounds in 50 cc. of the 
urine having been determined, it is subtracted from the weight of 
the total quantity of sulphur in 50 cc. of the urine; the remainder 
is the weight of sulphur of compounds other than sulphates and 
esters in 50 cc. of the urine. 

CHLORINE. 
VOLHARD'S METHOD, MODIFIED BY SALKOWSKI. 

The chloride and sulphocyanide of silver are insoluble in 
water and dilute nitric acid, while ferric sulphocyanide is soluble 
and imparts a red color to its solutions. Reactions by which 
these bodies are formed are understood by the equations : — 

NaCl -f AgN0 3 = NaN0 3 -f AgCl. 

AgN0 3 + NH 4 (CNS) = NH 4 NO s + Ag(CNS). 

Fe 2 (S0 4 ) 3 + 6NH 4 (CNS) = 3 (NH 4 ) 2 S0 4 + Fe 2 (CNS) 6 . 

A definite quantity of urine, having been rendered strongly 
acid with nitric acid, is treated with an excess of a solution of 
silver nitrate, the strength of which is known when the silver 
chloride is separated by filtering, and the quantity of silver in the 
filtrate determined by titrating with a standardized solution of 
ammon. sulphocyanide. From the quantity of silver in the solu- 
tion added, the quantity found in solution is subtracted; the 
remainder is the quantity of silver with which the chlorine in 
the urine has combined. If the titration with ammon. sulpho- 
cyanide be carried on without the separation of silver chloride, 



154 CHEMICAL ANALYSIS OF THE URINE. 

the results are unreliable, as the silver chloride reacts on silver 
sulphocyanide, causing the quantity of ammon. sulphocyanide 
to vary. In titrating with ammon. sulphocyanide, the ferric salt 
is the indicator. 

PREPARATION OF SOLUTIONS. 
SOLUTION OF SILVER NITRATE. 

Dissolve pure crystallized silver nitrate in distilled water, so 
that the strength of the solution is 29.075 grms. in 1000 cc. The 
quantity of water in which to dissolve a weighed quantity of 
silver nitrate is ascertained by the equation of 29.075 : 1000 : : 

No. of grms. 

AgN0 3 weighed : x or number of cc. water. The silver nitrate, 
when weighed, is put into a dry flask or bottle provided with a 
glass stopper, and the required amount of water is added. 1 cc. 
of the silver solution contains the quantity of silver which will 
combine with the chlorine of 0.0 1 grm. sodium chloride or 
0.00606 grm. chlorine. 

SOLUTION OF FERRIC ALUM. 

Ferric sulphate may be employed as well as ferric alum, but as 
the ammon. sulphate in the latter takes no part in the reaction, 
and as the compound is generally obtained free of chlorine, it is 
preferred. A saturated solution is employed, which is prepared 
by dissolving 50 grms. in 1000 cc. distilled water. 

SOLUTION OF AMMONIUM SULPHOCYANIDE. 

Dissolve about 7 grms. ammon. sulphocyanide in 11 00 cc. dis- 
tilled water, and having mixed well by shaking, fill a 25 cc. 
burette with the solution to the o mark. For the purpose of 
standardizing this solution with the solution of silver nitrate, 
10 cc. of the silver solution is introduced into a beaker and 
diluted with about 100 cc. water, and to the fluid 4 cc. nitric 
acid, sp. gr. 1.2, and 5 cc. of the solution of ferric alum are added ; 
and having mixed well with a stirring rod, the fluid is titrated 
with the solution of ammon. sulphocyanide, until, by the addi- 
tion of 0.1 cc, a red color or tint is imparted to the fluid, recog- 
nized by placing the beaker on a white background. Titrations 
are repeated until an agreement is reached, when 1000 cc. of the 
solution is diluted so that 25 cc. corresponds to 10 cc. of the 



TITRATION OF THE URINE. 155 

silver solution. For example: 21.3 cc. of the sulphocyanide 
solution was required to produce the red color with 10 cc. of the 
silver solution; the addition of 3.7 cc. water to 21.3 cc. of the solu- 
tion would be required to dilute the solution to 25 cc, and to 
dilute 1000 cc. of the solution 173.7 cc water would be required 
(21.3 : 3.7 : : 1000 : x or 173.7). 



TITRATION OF THE URINE. 

Into a 100 cc. graduated flask, provided with a glass stopper, 
introduce 10 cc. urine, 50 cc. water, 4 cc. nitric acid, sp. gr. 1.2, 
and 15 cc. of the solution of silver nitrate. Having closed the 
flask with the stopper, shake several minutes, and with a fine 
stream of water from a wash bottle rinse the end of the stopper 
and fill the flask with water to the mark. Close the flask again 
and mix well by shaking, and after settling, filter through a dry 
filter paper into a dry graduated 80 cc. flask. In filtering, the 
clear fluid is carefully decanted into the filter without agitating 
the precipitate. When the filtrate reaches the mark of 80 cc, the 
filter is removed, the filtrate introduced into a 300 cc. flask, and 
the flask rinsed with water several times and rinsings added to 
the filtrate. Add 5 cc. of the solution of ferric alum to the fluid 
and titrate with the solution of ammon. sulphocyanide from a 
burette, until, after having mixed well by agitating the flask, a 
permanent red color or tint is imparted to the fluid by the addi- 
tion of 0.1 cc of the solution. 

CALCULATION. 

In the fluid titrated there was introduced 15 cc of the solution 
of silver nitrate. If this quantity of silver solution were titrated 
with the solution of ammon. sulphocyanide in the absence of 
chlorides, 37.5 cc. of the sulphocyanide solution would be required, 
but chlorine of the urine having combined with some of the 
silver, less of the sulphocyanide solution would be required. 
The quantity of chlorine which has combined with silver is deter- 
mined by first estimating the quantity of silver in excess or that 
remaining in solution by titrating the filtrate with the solution of 
ammon. sulphocyanide and subtracting the number of cc. required 
from 37.5, the remainder is equivalent to a definite quantity of 



156 CHEMICAL ANALYSIS OF THE URINE. 

silver or chlorine. Example: 5.9 cc. of the solution of ammon. 
sulphocyanide was required for the silver in the filtrate which 
represents eight-tenths of the urine taken, therefore, for the filtrate 
and wash water of the 10 cc. urine, 7.37 cc. of the sulphocyanide 
solution would be required ( IO * 5 ' 9 = 7.37). As 15 cc. of the 
silver solution employee! requires 37.5 cc. (10 : 25 : : 15 : 37.5) 
of the ammon. sulphocyanide solution, and after coming in con- 
tact with the urine only 7.37 cc. was required ; therefore, chlorine 
equivalent to 30.1 cc. of the sulphocyanide solution combined 
with the silver, which is in 12 cc. of the solution of silver nitrate 
(37-5 : 1 5 : : 30. 13:1 2). That is, in 10 cc. urine there is suffi- 
cient chlorine to combine with the silver in 12 cc. of the standard- 
ized solution of silver nitrate, and as 1 cc. of the solution cor- 
responds to 0.00606 grm. chlorine, 12 cc. would correspond to 
0.07272 grm. There being this quantity of chlorine in 10 cc. 
of the urine, in 100 cc. there would be 0.7272 grm. chlorine. 

NEUBAUER'S METHOD. 

By Neubauer's method the quantity of the chlorine in urine is 
estimated by titrating with a solution of silver nitrate of known 
strength ; the indicator employed is a solution of the yellow 
chromate of potassium. The chlorine having combined with 
silver, and with a slight excess of silver solution silver chromate 
is formed, which imparts a red color or tint to the mixture, but 
before titrating the organic matter of the urine is oxidized. For 
this purpose introduce 10 cc. of the urine into a platinum dish or 
crucible, to which add 1 grm. sodium carbonate and about 3 
grms. potassium nitrate — both free of chlorides — and evaporate 
to dryness on a water bath. When dry, place the dish or cru- 
cible on a triangle support, and with a small flame of a gas or 
spirit lamp heat gradually by keeping the flame in lateral move- 
ment, so that the heat be applied equally. Increase the heat 
until the residue melts, forming a white mass when the lamp is 
removed. Dissolve the mass, when cold, in water, and empty 
into a 150 cc. flask. Before transferring the fluid, place a small 
funnel in the neck of the flask, so as to prevent loss. Rinse the 
dish and funnel and add the rinsings to the fluid, neutralize the 
solution by first rendering distinctly acid with dilute nitric acid 
and neutralizing; with a solution of sodium carbonate. To the 



POTASSIUM AND SODIUM. 157 

neutral solution add two or three drops of a saturated solution 
of the yellow chromate of potassium, when it is prepared for 
titrating. The solution of silver nitrate of the strength employed 
in Volhard's method of estimating chlorine in the urine is pre- 
ferred. For the preparation of this solution, refer above. The 
neutralized solution is titrated with the solution of silver nitrate 
from a burette until, after mixing well by agitating the flask, the 
mixture is changed to a red color or tint by the addition of o.i cc. 

CALCULATION. 

For the weight of chlorine or sodium chloride in grms., mul- 
tiply the number of cc. of the silver solution employed in the 
titration by 0.00606 for chlorine, and by 0.0 1 for NaCl ; the pro- 
duct of each multiplied by 10 will give the number of grms. in 
100 cc. urine. 

REMARKS. 

In Volhard's method of estimating chlorine in the urine, potas- 
sium sulphocyanide may be employed instead of the ammonium 
salt. The solution of either salt undergoes no change by keep- 
ing in well filled glass-stoppered bottles, in a cool, dark place. If 
the nitric acid, sp. gr. 1.2, is colored, heat in a flask on a water 
bath until the nitrous fumes are driven off. 

The most difficult part of Neubauer's method is to neutralize 
the solution of the fused mass. If, however, too much of the 
solution of sodium carbonate be added after having rendered 
acid with nitric acid, the solution may be rendered slightly acid 
with acetic acid, or the chlorine may be estimated by Volhard's 
method. With this in view, render the solution strongly acid 
with nitric acid, add 15 cc. of the silver solution, boil 15 to 20 
minutes, to drive off the nitrous acid, and proceed as with urine 
treated with acid and silver solution. 



POTASSIUM AND SODIUM. 

In the estimation of potassium and sodium in the urine, chlo- 
rides of these metals are weighed ; but before this, the phosphoric 
and sulphuric acids are separated, the organic matter oxidized, 
and the calcium magnesium and also barium employed in sepa- 
rating the acids are removed. 



158 CHEMICAL ANALYSIS OF THE URINE. 

50 cc. of the urine is mixed with 50 cc. barium mixture* in a 
dry beaker. Having stood three or four hours, filter into a dry, 
graduated 100 cc. glass cylinder, and when the nitrate reaches 
the mark of 80 cc, remove the filter. Evaporate the filtrate 
80 cc. with the rinsings of the cylinder in a platinum dish to dry- 
ness on a water bath, and when evaporated, transfer the dish to a 
triangle support, and heat gently with a small flame. If not heated 
gradually, by the sudden evolution of steam or by the rapid 
oxidation of organic matter, loss may result. A high temperature 
is avoided, as the chlorides volatilize at a white heat. When cold, 
treat the residue in the dish with 20 cc. water and render the 
solution acid with dilute hydrochloric acid, then the fluid is ren- 
dered alkaline with ammonium hydrate and a solution of ammo- 
nium carbonate added until a precipitate ceases to form. The 
metals of the alkaline earths separate as carbonates. Filter, and 
wash the precipitate with water and add the wash water to 
the filtrate. Evaporate the filtrate and wash water in the plati- 
num dish to dryness on a water bath, and heat the residue with 
great care, to avoid crepitation of the chlorides. Repeat the 
process of dissolving in a small quantity of water, rendering 
alkaline with ammon. hydrate, adding a small quantity of a solu- 
tion of ammon. carbonate, filtering, washing, evaporating and 
heating the residue until ammon. carbonate ceases to produce a 
precipitate, when the residue is heated in a weighed platinum 
dish provided with a cover, and having cooled in a desiccator, it 
is weighed. The difference in the weight of the empty dish and 
the weight of the dish + residue is the weight of potassium and 
sodium chlorides in 40 cc. of the urine. The chlorides in the dish 
are dissolved in a small quantity of water, and the solution trans- 
ferred to a porcelain dish and the dish rinsed with water, the 
rinsings added to the solution in the porcelain dish. To the solu- 
tion add an excess of a solution of platinum chloride (1 part PtCl 4 
and 10 parts water) and evaporate on a water bath, but the evapo- 
ration should not be carried so far that water of crystallization of 
compounds is driven off. Treat the residue with a few drops of 
the platinum water to dissolve the sodium compound, and. add a 



* Composed of 2 vols, cold saturated solution of pure barium hydrate and I vol. 
cold saturated solution of barium chloride. 



AMMONIA. 159 

mixture of 4 vols, absolute alcohol and 1 vol. ether; with a glass 
rod stir several minutes, let stand in a cool place 30 minutes, and 
filter through a small, dry filter paper and wash well with the 
mixture of alcohol and ether. When washed, dissolve the yellow 
precipitate on the paper with hot water, and, solution having taken 
place, wash with hot water until the platinum compound is 
washed from the paper. Evaporate the filtrate and wash water 
in a weighed platinum dish provided with a cover on a water 
bath, and dry the residue at ioo° C. in an air or steam bath, and 
after cooling in a desiccator, it is weighed. 

CALCULATION. 

Multiply the weight of the platinic potassium chloride by 
0.3055 ; the product is the weight of potassium chloride in the 
urine. Having determined the weight of potassium and sodium 
chlorides, the weight of potassium chloride is subtracted from it. 
the remainder is the weight of sodium chloride. To determine 
the weight of potassium, multiply the weight of potassium 
chloride by 0.5246. The weight of sodium is determined by 
multiplying the weight of sodium chloride by 0.3938. As the 
quantities are of 40 cc. urine, multiply by 2.5 ; the product is the 
weight of each in 100 cc. of the urine. 

AMMONIA. 
SCHLOSING AND NEUBAUER'S METHOD. 

When a salt of ammonium in solution comes in contact with 
calcium hydrate, ammonia gas is set free ( (NH 4 ) 2 C0 3 + Ca(OH) 2 
= CaCO.3 + 2H 2 — 2NH3). When ammonia comes in contact 
with dilute sulphuric acid, ammon. sulphate is formed (2NH3 -{- 
H 2 S0 4 = (NH 4 ) 2 S0 4 ). The ammonia, in a certain volume of 
urine, is liberated by the action of calcium hydrate, and is 
absorbed by dilute sulphuric acid of known strength, and by 
titrating with one-fifth normal potassium hydrate the quantity of 
ammonia combined with the acid is determined. For the prepa- 
ration of normal sulphuric acid, refer to page 1 12, and for the pre- 
paration of normal potassium hydrate, refer to page 1 09. From the 
normal solutions the one-fifth of each is prepared by introducing 
100 cc. into a 500 cc. graduated flask and filling with water to 
the mark. Before using, the solutions are mixed by shaking. A 




160 CHEMICAL ANALYSIS OF THE URINE. 

saturated solution of calcium hydrate and a solution of litmus 
are also required. Calcium hydrate is mixed with water, and 
after standing one or two hours and frequently shaking, the solu- 
tion is filtered and kept in a well- 
stoppered bottle. For the prepara- 
tion of a solution of litmus paper,, 
refer to page 1 14. Besides these 
solutions, the apparatus represented 
°y Fig. 35 is required. The appa- 
ratus comprises a bell jar or receiver 
with ground margin, a glass plate 
with ground surface, on which the 
receiver is placed to prevent the 
escape of ammonia, two evaporating dishes and a tripod. The 
evaporating dishes should be shallow. The internal (transverse) 
diameter of the bell jar is 14 cm. 

30 cc. of the one-fifth normal sulphuric acid is introduced into 
one of the evaporating dishes, and a sufficient quantity of the 
solution of litmus is added to impart a distinct red color to the 
fluid. Into the other evaporating dish introduce 20 cc. of clear 
or filtered urine and 20 cc. of the saturated solution of calcium 
hydrate. The dish is then placed on the glass plate under the 
tripod supporting the other dish containing the dilute acid, when, 
with some grease, the receiver is fixed in place. After standing 
three days in a cool place where the temperature is not subject 
to variation, empty the dish containing the dilute acid into a 
beaker, and with water rinse the dish several times, adding the 
rinsings to the fluid in the beaker. With the one-fifth normal 
potassium hydrate in a burette, titrate the acid solution until, 
after stirring with a glass rod, the fluid turns purple or blue by 
the addition of 0.1 cc. 

CALCULATION. 

In the absence of ammonia, 30 cc. of the one-fifth normal acid 
would require 30 cc. of the one-fifth normal potassium hydrate, 
but by the absorption of ammonia some of the acid is neutralized, 
and, therefore, less of the potassium hydrate is required. If, for 
example, 27.5 cc. of the one-fifth normal potassium hydrate be 
required to neutralize the 30 cc. one-fifth normal sulphuric acid 
employed, 2.5 cc. of the acid is in combination with ammonia. 



CALCIUM OXIDE. 161 

I cc. of the one-fifth normal sulphuric acid will combine with 
0.0034 grm. ammonia and 2.5 cc. of the acid corresponds to 
0.0085 grm. ammonia (2.5 X 0.0034 = 0.0085). There being 
0.0085 grm. ammonia in 30 cc. urine, in 100 cc. there is 0.02833 

C-^^- = 0.02833.) 



CALCIUM AND MAGNESIUM OXIDES. 

These bodies in the urine are estimated by the ordinary gravi- 
metric methods employed in estimating them in inorganic com- 
pounds ; besides, they are estimated by volumetric processes. 
Calcium is precipitated as an oxalate with a solution of ammon. 
oxalate in the presence of magnesium and phosphoric acid, if 
the solution be rendered alkaline with ammon. hydrate and then 
acidified with acetic acid. Calcium is separated from the urine 
in the form of an oxalate in the volumetric as well as in the 
gravimetric method. The magnesium is precipitated from an 
acid solution in the presence of a soluble phosphate by ren- 
dering the solution alkaline with ammon. hydrate as magnesium 
ammon. phosphate (MgNH 4 P0 4 ). After the separation of calcium 
from the urine by means of ammon. oxalate, the urine contains 
all of the reagents necessary, when rendered alkaline with ammon. 
hydrate, to form the insoluble magnesium ammon. phosphate, 
but the precipitation is facilitated by the presence of ammon. 
hydrogen phosphate. The magnesium is precipitated in this 
form in both the gravimetric and volumetric methods. 

CALCIUM OXIDE. 
THE GRAVIMETRIC METHOD. 

250 cc. clear or filtered urine is introduced into a beaker and 
treated with ammon. hydrate until a precipitate is formed, when 
it is rendered acid with acetic acid. The phosphates of calcium 
and magnesium, which precipitate by rendering the urine alka- 
line, are dissolved by the acetic acid. Place the beaker on a 
wire gauze and heat until the fluid begins to boil, when the cal- 
cium is precipitated by adding an excess of a saturated solution 
of ammon. oxalate. Mix well by stirring with a glass rod, 
avoiding contact with the surfaces of the beaker. Place the 
beaker on a water bath and let remain until the precipitate has 



162 CHEMICAL ANALYSIS OF THE URINE. 

settled. Introduce I or 2 cc. of the ammon. oxalate solution into 
the clear fluid from a pipette; if no precipitate is produced a suffi- 
cient quantity has been added. Filter through a filter paper free 
of ash, and collect the precipitate by means of a glass rod, over 
the end of which a short piece of rubber tubing is placed. Wash 
the precipitate with distilled water until the wash water remains 
clear when tested with a solution of silver nitrate and a few 
drops of nitric acid. Preserve the filtrate and wash water for the 
estimation of magnesium oxide. Dry the filter with precipitate 
at 100 C, and when dry, place into a weighed platinum crucible 
and heat with a gas or spirit lamp in the open crucible. The 
filter having been ashed, place the lid on the crucible and heat 
with the blast lamp at a high temperature 10 or 15 minutes, and 
the crucible having cooled in a desiccator, it is weighed. The 
process of heating and weighing should be repeated until the 
weight becomes constant. The difference between the weight of 
the empty crucible and that of the crucible and contents is the 
weight of calcium oxide in 250 cc. of the urine. By employing 

Wt. of 

the equation 250 : CaO : : 100 : x, the weight of calcium oxide in 
100 cc. is determined. 

THE VOLUMETRIC METHOD^NEUBAUER. 

By this method the calcium is separated from the urine, as in 
the gravimetric method, and by heat it is transformed into the 
oxide, when it is treated with an excess of dilute hydrochloric 
acid of known strength, after which the quantity of free hydro- 
chloric acid is estimated by titrating with a solution of potassium 
hydrate, and by calculation the quantity of calcium oxide is 
determined. One-half normal hydrochloric acid and potassium 
hydrate are employed. For the preparation of normal hydro- 
chloric acid, refer to page 113, and for the preparation of normal 
potassium hydrate, refer to page 109. A one-half normal solu- 
tion is prepared by mixing one volume of the normal solution 
with one volume distilled water. This is accomplished without 
difficulty by introducing 250 cc. of the normal acid or potassium 
hydrate into a 500 cc. graduated flask and filling with water to 
the mark. The calcium in 250 cc. urine having been precipitated 
in the form of oxalate, as in the gravimetric method given above, 
and the precipitate having been washed, dried, the filter ashed 



MAGNESIUM OXIDE. 163 

and the oxalate decomposed by heat into the oxide, the latter is 
transferred from the crucible to a 200 cc. flask by means of a 
hair pencil, when to the oxide 50 cc. water is added, and after 
mixing well by agitating the flask, 10 cc. of the one-half normal 
hydrochloric acid is introduced into the mixture from a burette. 
Calcium oxide will dissolve in the acid, and by heating the solu- 
tion any carbonic acid present is driven off. Add a sufficient 
quantity of the solution of litmus to impart a distinct red color 
to the fluid. From a burette containing the one- half normal 
potassium hydrate, titrate the fluid until, after mixing well by 
agitating the flask, the color of the solution changes purple or 
blue by the addition of 0.1 cc. 

CALCULATION. 

10 cc. of the one-half normal potassium hydrate would be 
required to neutralize the 10 cc. one-half normal hydrochloric 
acid if no base were present, but as calcium oxide combines with 
the acid, less of the potassium hydrate would be required. If, 
for example, 7.5 cc. of the potassium hydrate solution neutral- 
ized the acid, 2.5 cc. of the acid was absorbed by the calcium 
oxide, and as 1 cc. of the half normal hydrochloric acid will 
combine with 0.014 grm. calcium oxide, therefore, there is in 
250 cc. of the urine 0.035 g rm - calcium oxide (0.014 X 2.5 == 
0.035), an d in 100 cc. 0.014 grm. 

MAGNESIUM OXIDE. 
GRAVIMETRIC METHOD. 

For the estimation of the quantity of magnesium oxide, the 
filtrate and wash water of the calcium oxalate from 250 cc. urine 
are employed. Refer above to gravimetric method, page 161. 
Evaporate the fluid in an evaporating dish to about 200 cc, 
transfer to a beaker, rinse the dish with water and add rinsings 
to the fluid in the beaker. To the fluid add one-fourth its volume 
strong ammonium hydrate and 5 or 10 cc. of a clear solution 
of sodium ammon. hydrogen phosphate (1 part of the salt and 
10 parts water), the magnesium is precipitated as magnesium 
ammon. phosphate. Having mixed well with a stirring rod 
without the rod coming in contact with the beaker, let stand in a 
warm place, protected from dust, 12 to 24 hours, when the solu- 



16 i CHEMICAL ANALYSIS OF THE URINE. 

tion is filtered through a filter paper free of ash. Collect the 
precipitate by means of a glass rod provided with a short piece 
of rubber tubing placed over its end, and wash the precipitate 
on the filter with a mixture of 2 vols, water and I vol. 
ammon. hydrate until the wash water yields no precipitate, 
after having been boiled to drive off ammonia acidified with 
nitric acid and treated with a solution of silver nitrate. Dry the 
filter with precipitate at ioo c C. and transfer from the paper to a 
clean, dry watch glass as much of the precipitate as is possible, 
and place the filter in a weighed platinum crucible, and ash it by 
heating the crucible with a gas or spirit lamp to redness. When 
the ash has become white or gray, remove the lamp and transfer 
the precipitate from the watch glass to the crucible by means of 
a fine hair pencil, and with the crucible closed with the lid heat 
gradually to redness. If the precipitate, after having been heated, 
is not white, it is due to the presence of carbon of organic 
matter, to oxidize which, introduce a crystal of pure ammon. 
nitrate into the crucible and heat again, or oxidation will take 
place by heating in the open crucible to redness 30 minutes. 
Having cooled in a desiccator, weigh. The body weighed is mag- 
nesium pyrophosphate ; the magnesium ammon. phosphate having 
been decomposed by heat forming the pyrophosphate, as shown 
by the equation, 2MgNH 4 P0 4 = Mg 2 P 2 7 + H 2 + 2NH3. 

CALCULATION. 

Multiply the weight of the precipitate by 0.36036; the product 
of which is the weight of magnesium oxide in 250 cc. of the 
urine, and two-fifths of which is the quantity in 100 cc. urine. 

THE VOLUMETRIC METHOD— NEUBAUER. 

The calcium having been separated from 250 cc. urine, as by 
the gravimetric method of estimating calcium, page 161, and the 
magnesium precipitated as magnesium ammon. phosphate from 
the filtrate and wash water as by the gravimetric method of esti- 
mating magnesium^ page 163, and the precipitate having been 
well washed, it is transferred to a beaker by perforating the 
filter with a glass rod and by means of a fine stream of water 
from a wash bottle. When the precipitate is removed from the 
paper, it is dissolved with dilute acetic acid, and the phosphoric 



MAGNESIUM OXIDE. 165 

acid in the solution is determined by titrating with a standardized 
solution of uranium acetate. For the preparation of the uranium 
solution and the method of titrating, refer to pages 144 and 145. 

CALCULATION. 

Multiply the weight of phosphoric acid, P 2 5 , found by 0.5633 ; 
the product of which is the weight of magnesium oxide in 250 cc. 
of the urine, and two-fifths of which is the quantity in 100 cc. 
urine. 



CHAPTER XII. 

Albumen, Scherer's Method — Globuline, Hammarsten's Method — Pohl's Method — 
Hemialbumose, Gravimetric and Optical Methods — Peptone, Optical Method — 
Remarks on the Estimation of Albuminous Bodies — Sugar, Fehling's Method — 
Fehling's Method, Modified by Pavy — Roberts' Method — Optical Method — Re- 
marks on the Estimation of Sugar in the Urine. 



ALBUMEN. 
SCHERER'S METHOD. 

Urine in which albumen is to be estimated, if not clear, is 
filtered. Into a beaker of about 200 cc. capacity, 100 cc. urine is 
introduced. If the reaction of the urine is not strongly acid, add 
acetic acid until the reaction is decidedly acid, but avoid an 
excess of the acid. Suspend the beaker in a water bath and 
keep the water in the bath at> the boiling temperature. At the 
expiration of 30 minutes, if, by transmitted light, the urine is 
clear between the flakes of coagulated albumen, the precipita- 
tion is complete. If, however, the urine is cloudy, a small quan- 
tity of acetic acid is added, the urine stirred, and the heat con- 
tinued, when the separation of albumen in flakes will take place. 
Filter through a filter, having been dried at no° C. between 
watch glasses and cooled in a desiccator and weighed. The albu- 
men having been transferred to the filter, is washed with water. 
As the filtering and washing are likely to require several hours, 
a filter pump or aspirator bottle may be employed with advantage, 
the filter having the support of a platinum cone. The washing is 
continued until no cloudiness is produced when tested with a 
solution of silver nitrate and some nitric acid. Having been 
washed with water, wash with about 50 cc. absolute alcohol, fol- 
lowed by about the same quantity of ether. Any fat present 
is removed by the alcohol and ether, and the water is so far 
removed as to facilitate the drying. The funnel is covered with 
paper or a glass plate and placed upright in an air bath and 
heated gradually until the paper and precipitate are somewhat dry, 
when the filter, with the precipitate, is placed between the watch 
glasses employed before. The heating in the air bath at 1 io° C. 

166 



GLOBULINE. 107 

is continued until the weight becomes constant, which is ascer- 
tained by heating two hours, cooling in a desiccator and weigh- 
ing, repeating the process until the weight becomes constant. 
The difference in weight caused by the precipitate is taken as the 
weight of albumen, except in case the urine contains much albu- 
men ; when the filter paper and precipitate are ashed and the ash 
weighed in a platinum crucible. By subtracting the weight of 
the ash from that of the precipitate, the remainder is the weight 
of albumen, or, instead of ashing, 50 cc. urine may be taken and 
50 cc. water added before acidifying and heating. The albumen, 
when dry, should not exceed 0.3 grm. in weight; if less, the 
quantity of inorganic matter present is very small. 



GLOBULINE. 
HAMMARSTEN'S METHOD. 

For outline of the method for separating globuline from the 
urine, refer to page 47. In separating globuline from albuminous 
urine with magnesium sulphate, the urine is rendered as nearly 
neutral as possible without precipitating calcium and magnesium 
phosphates. For this purpose dilute acetic acid and a weak 
solution of sodium carbonate are used. If the urine is highly 
colored and the globuline not in very small quantity, it is diluted 
with water 25 or 50 cc. urine with 75 or 50 cc. water. If the 
quantity of globuline is too small to admit of dilution, and the 
urine is highly colored, surround the flask or beaker containing 
it with ice broken in small fragments, and when the temperature 
of the urine has been reduced to 2° or 3 C. several hours, the 
urates may precipitate, carrying with them much of the coloring 
matter of the urine. Filter, while cold, through a dry folded 
filter paper. To 100 cc. urine or urine and water, if diluted in a 
beaker, add 80 grms. pure powdered magnesium sulphate. Stir 
continually until the urine becomes saturated ; avoid, however, 
the formation of foam. Add more of the magnesium sulphate, 
if necessary, and let stand 24 hours, stirring occasionally. Globu- 
line precipitates while' serum-albumen, hemialbumose and pep- 
tone remain in solution. Filter through a small filter paper free 
of ash, having been dried at 110 C. in an air bath between two 
watch glasses, and weighed after having cooled in a desiccator. 



168 CHEMICAL ANALYSIS OF THE URINE. 

Before filtering, the filter is wet with a saturated solution of mag- 
nesium sulphate. Wash the precipitate with a saturated solution 
of magnesium sulphate until the wash water, when acidified with 
acetic acid, remains clear by boiling (absence of albumen), when 
the funnel is placed upright in an air bath — covered to protect 
from dust — and heated to uo° C. at least 3 hours. By the heat 
the globuline becomes insoluble. Let cool and wash with hot 
water until the wash water, when tested with a solution of barium 
sulphate, produces no cloudiness (absence of MgS0 4 ), after 
which the precipitate is washed with absolute alcohol two or 
three times, and as many times with ether. Place the filter and 
precipitate between the watch glasses. Dry at uo° C, and after 
cooling in a desiccator, weigh. Repeat the process until the 
weight becomes constant. The increase in weight caused by 
the precipitate having been noted, the filter and precipitate are 
ashed in a weighed platinum crucible, and the weight of the ash 
is subtracted from the weight of the dry precipitate ; the remain- 
der is the weight of globuline in the quantity of urine employed. 

POHL'S METHOD. 

Render the urine neutral by employing dilute acetic acid and a 
weak solution of ammonium hydrate ; filter, if necessary, and mix 
in a beaker 100 cc. of the filtrate — or urine, if not filtered — with 
100 cc. of a saturated solution of neutral ammonium sulphate, 
and after standing one hour, separate the precipitate — globuline 
— by filtering through a weighed filter, having been dried at 
110 C, and wash the precipitate with a mixture of 1 volume 
saturated solution of ammon. sulphate and 1 volume water, until 
the wash water yields no precipitate or cloudiness when tested 
with a solution of potassium ferrocyanide acidified with acetic 
acid. Place the funnel upright in an air bath and heat three 
hours at no° C. to render the globuline insoluble in water. 
When cool, wash with hot water until the wash water is free of 
ammon. sulphate, known by yielding no turbidity when tested 
with a solution of barium chloride. The washing is then con- 
tinued with 100 cc. alcohol, followed by an equal volume of 
ether. Dry the filter and precipitate between the watch glasses, 
employed before at no° C, and having cooled in a desiccator, 
weigh. This process is repeated until the weight is constant. 



PEPTONE. 169 

Having ascertained the weight of the precipitate, ash it with the 
filter in a weighed platinum crucible, and subtract the weight of 
the ash from the weight of the precipitate, the remainder is the 
weight of globuline in ioo cc. urine. 

HEMIALBUMOSE. 

There is no method for determining the exact quantity of 
hemialbumose in urine, but the quantity within narrow limits is 
determined. To prepare urine for the estimation, separate the 
albumen by adding to 500 cc. urine yi its volume (85 cc.) of a 
saturated solution of common salt — sodium chloride — and if the 
urine is not decidedly acid in reaction, dilute acetic acid is added 
until it is acid. Heat to the boiling temperature and filter. To 
precipitate hemialbumose in the filtrate add an equal volume of 
a saturated solution of common salt, and after stirring filter 
through a dry weighed filter paper, free of ash, and having brought 
the precipitate on the paper, wash it with a saturated solution of 
common salt. 

Dry the filter with precipitate at no° C. until the weight is 
constant, and note the increase in weight caused by the pre- 
cipitate and some common salt. Ash the filter and precipitate, 
and weigh the ash in a porcelain or platinum crucible, and 
subtract the weight of the ash from the weight of the precipitate : 
the remainder is the approximate weight of hemialbumose in 500 
cc. urine. 

Instead of weighing the precipitate, it may, without drying, be 
dissolved in water, and the quantity determined, according to 
Salkowski, by means of a polariscope. The specific rotation of 
hemialbumose is — 75 °. For the use of the polariscope, refer 
below. By employing a tube 10 cm. long, the equation for 

degrees 

the percentage is — 75 : 100 : : rea d : x. If the solution of hemi- 
albumose examined occupies the volume of 500 cc, no further 
calculation is required, but if 250 cc, then the per cent, found is 
divided by 2: 

PEPTONE. 
HOFMEISTER'S METHOD. 

This method is based on the property peptone has, when in 
solution, of changing the plane of vibration of a ray of light 
having been polarized. For outline of description of polariscope > 



170 CHEMICAL ANALYSIS OF THE URINE. 

refer below. To prepare the urine for examination, it is decol- 
orized by treating 80 cc. of the urine in a 100 cc. graduated 
flask with a few drops of a solution of neutral lead acetate, when 
the flask is filled with water to the mark. Shake well and filter 
through a dry filter paper into a dry flask, and examine the 
filtrate with a polariscope. The specific rotation of peptone is 
— 63.5 °. The calculation is made by the equation 63.5 : 100 : : 

degrees 

read : x, if a tube 10 cm. long be employed. From the per cent, 
or weight of peptone found in 100 cc. of the solution examined, 
the quantity in 100 cc. urine is determined by multiplying by \ or 
1.25, as 100 cc. of the solution corresponds to 80 cc. urine. This 
method does not provide for the presence of albumen or hemi- 
albumose in the urine, as these bodies also change the plane of 
vibration of polarized light. If present they are separated (Hof- 
meister), by adding to 80 cc. acid urine a solution of sodium 
carbonate, until the urine is slightly acid in reaction, and in case 
the urine is alkaline dilute acetic acid is added until the urine is 
acid, yet avoiding an excess. The urine is then treated with 10 
cc. of a saturated solution of sodium acetate, when a solution of 
ferric chloride is added until the urine becomes somewhat red in 
color. Boil a short time, let cool, and with water dilute to the 
mark 100 cc, and having mixed well by shaking, filter through a 
dry filter into a dry flask or beaker. The filtrate when exam- 
ined with the polariscope should not yield a precipitate with a 
solution of potassium ferrocyanide and acetic acid. 

REMARKS. 

Of the great number of methods for the estimation of albumen 
in the urine, however ingenious some of them are, there are 
none which lead to more accurate results than that of Scherer, 
although the drying and filtering by this method require time 
and close attention. The methods of estimating hemialbumose 
and peptone are not exact. 



SUGAR. 

FEHLING'S METHOD. 

For the facts on which this method is based, refer to page 
50. 



PREPARATION OF SOLUTIONS. 171 

PREPARATION OF SOLUTIONS. 
SOLUTION OF COPPER SULPHATE. 

Commercial copper sulphate is purified by dissolving as much 
of the salt in hot water as possible, and filtering while hot into 
a beaker, and let crystallize by standing in a cool place. The 
mother liquid is poured off and the crystals collected in a funnel 
in the neck of which there is some spun glass, to prevent the 
crystals from passing through when they are washed with a 
small quantity of cold water. Dissolve again in hot water and 
continue as before. The crystals are dried by pressing several 
times between porous paper. To prepare the solution, weigh on 
a balance of precision 34.639 grms. of the copper sulphate. Intro- 
duce the salt into a 500 cc. graduated flask and dissolve in some 
water and fill with water to the mark. Mix well by shaking. 

SOLUTION OF SODIUM POTASSIUM TARTRATE AND SODIUM HYDRATE. 

Dissolve 173 grms. sodium potassium tartrate in about 300 cc. 
warm water in a 500 cc. graduated flask. If the solution is not 
clear, filter, and to the solution add 50 grms. sodium hydrate, and 
when dissolved fill with water to the mark, but not until the 
temperature is reduced to about iy° C. 

FEHLING'S SOLUTION. 

Fehling's solution is prepared by mixing an equal volume 
of the solution of copper sulphate and the solution of sodium 
potassium tartrate and sodium hydrate; but as Fehling's solution 
is liable to change, the solutions should be prepared for immediate 
use and, therefore, in less quantities. 

Before using Fehling's solution, test it by heating to the boil- 
ing temperature in a flask 10 cc. of the solution diluted with 
about 40 cc. water, and after cooling and standing several 
minutes, if no Cu 2 is found remaining on the bottom of the 
beaker when the solution is decanted, there is no reducible 
substance in the solution. If Fehling's solution has been pre- 
pared with care, there' is no necessity of testing its strength with 
a solution of grape sugar. 1 cc. of Fehling's solution == 5 milligr. 
diabetic or grape sugar. 



172 CHEMICAL ANALYSIS OF THE URINE. 

THE TITRATION. 
The titration is made with the urine instead of the reagent, 
io cc. Fehling's solution is introduced into a 100 cc. flask with 
a pipette. For accurate results, sugar in the urine should 
be in such quantity that between 6 and io cc. of the urine 
would be required to reduce the CuO in io cc. of Fehling's 
solution, but as there is generally more sugar in diabetic urine, it 
requires dilution with water. Urine containing but little coloring 
matter, and by qualitative tests yields reactions indicating the 
presence of much sugar, may be diluted from i to io volumes 
without a preliminary estimation. On the other hand, if the 
indications are that the amount of sugar is not great, preliminary 
estimations are made. A burette having been cleaned and 
dried, is filled with urine, and the io cc. Fehling's solution in the 
flask is diluted with 40 cc. water placed on a wire gauze and 
heated with a gas or spirit lamp until it begins to boil, when 
0.5 cc. of the urine is added from the burette, a yellow or red 
precipitate will form. After shaking, let the precipitate settle. If 
sufficient urine has been added to reduce the CuO, the blue color 
of the solution will have disappeared, as seen by the surface of 
the fluid, when at an angle of about 45 °, with a white back- 
ground. If the blue color is perceptible, heat again to the boiling 
point, and add 0.5 cc. urine, and if the blue color still remains 
after settling, repeat the titrations until the blue color disappears. 
Having made one or two preliminary titrations, dilute the urine if 
necessary. In diluting use round numbers and measure in a 
graduated 100 cc. cylinder; for example, 3 cc. urine decolorizes 
10 cc. of Fehling's solution. Into the cylinder introduce 30 cc. 
of the urine and with water fill to the mark of 90 cc. ; the dilu- 
tion is 1 to 3. Theoretically, 10 cc. of Fehling's solution would 
require 9 cc. of the diluted urine. Titrations may now be made, 
adding 8.6, 8.8 and 9 cc. of the diluted urine, and if the cupric 
oxide is not all reduced, titrations are made, adding an increased 
quantity of 0.2 cc. until complete reduction takes place. After 
diluting the urine, time is saved by heating three or four flasks 
containing Fehling's solution with water at the same time, and by 
employing a 50 cc. burette containing the diluted urine, and 
keeping a record of the quantity of urine added to the contents of 
each flask. 



THE TITRATION. 173 

Instead of depending on the disappearance of the blue color of 
Fehling's solution, after settling, a small quantity of the fluid is 
decanted into a test tube and heated with o.i or 0.2 cc. of the 
urine, and if CuO is present a yellow or red precipitate will form, 
or, as has been recommended, a small quantity of the fluid may 
be filtered, the filtrate acidified with acetic acid and tested with a. 
solution of potassium ferrocyanide ; if CuO is present, a red pre- 
cipitate or coloration takes place, but by boiling urine with sodium 
hydrate ammonia is formed, which dissolves a small quantity of 
cuprous oxide, and by exposure to the air during the process 
of filtering it oxidizes, forming cupric oxide ; consequently, this 
method of determining when reduction of CuO has taken place is 
not satisfactory. 

CALCULATION. 

Suppose the urine was diluted from 1 to 9 and that 8.1 cc, of 
the diluted urine was required to reduce the copper oxide in 10 
cc. Fehling's solution. The urine in 8.1 cc. of the diluted urine 
is J of 8.1, which is 0.9 cc, hence there is in this amount of urine 
sufficient sugar to reduce the cupric oxide in 10 cc. Fehling's 
solution. 1 cc. Fehling's solution corresponds to 0.005 grm. 
sugar, and 10 cc. to 0.050 grm., therefore in 0.9 cc. urine 
there is 0.050 grm. sugar, and in 100 cc. urine 5.555 grms. sugar 
(0.9 : 0.050 : : 100 : x). 

FEHLING'S METHOD MODIFIED BY PAVY. 

In the estimation of sugar in the urine by Fehling's method, it 
is often very difficult to determine the point, in titrating, when the 
cupric oxide is reduced, as cuprous oxide remains suspended. To 
obviate this difficulty, Pavy's solution may be employed and the 
titration carried on in the absence of air. In this solution there 
is ammon. hydrate, by which the cuprous oxide or hydrate is held 
in solution, so that complete reduction of the cupric oxide is known 
by the solution losing its blue color. As cuprous oxide oxidizes 
readily by exposure to air, especially when dissolved in water by 
means of ammonia, provision is made for the exclusion of air 
while the reduction takes place. Pavy's solution has xV the 
reducing effect of Fehling's solution. 



174 CHEMICAL ANALYSIS OF THE URINE. 

PAVY'S SOLUTION. 
Introduce 12OCC. of Fehling's solution (for the preparation of 
which refer to page 171) into a 1000 cc. graduated flask contain- 
ing 300 cc. ammon. hydrate, sp. gr. 0.80; when 140 grms. sodium 
hydrate is added, and when solution has taken place and the 
temperature reduced to about iy° C, fill with distilled water to 
the mark, mix well by shaking. 

THE TITRATION. 
To prevent the access of air to the fluid titrated, a 200 cc. round 
bottom flask is provided with a cork having two holes, through 
one of which passes the dropper of the burette containing the 
urine, and through the other hole a bent glass tube passes which 
is connected with a chlor-calcium tube, filled with fragments of 
pumice saturated with dilute sulphuric acid. Introduce 50 cc. 
Pavy's solution into the round bottom flask, and when the solu- 
tion is heated to near the boiling point, urine is added from the 
burette, but, as reduction takes place more slowly than with 
Fehling's solution, care is taken not to boil the solution actively, 
or the ammonia may be driven off, the cuprous oxide precipitated 
and the solution lose much of its blue color before the cupric 
oxide has been completely reduced. The cupric oxide is reduced 
when the solution becomes colorless or slightly yellow in color. 
The difficulty encountered by the escape of ammonia before 
reduction takes place 'is obviated by introducing the dropper 
of a separating funnel into the flask through a hole in the stopper, 
and during the titration adding small quantities ammon. hydrate; 
but this precaution will not be found necessary if ammon. hydrate 
of the sp. gr. 0.80 be employed in preparing the solution, and 
nearly as much urine as is required for complete reduction of the 
CuO be added at once and the solution heated a few moments to 
the boiling point. In the employment of Pavy's solution urine is 
diluted if necessary, so that 7 to 10 cc. urine contains sugar in 
quantity to reduce the cupric oxide in 50 cc. Pavy's solution. 

CALCULATION. 

The quantity of sugar to reduce the CuO in 50 cc. Pavy's solu- 
tion is 0.025 grm., taking which into account the calculation is 
the same as by the employment of Fehling's solution. 



THE OPTICAL METHOD. 175 

ROBERTS' METHOD. 

This method rests on the fact that by fermentation of sugar in 
the urine the specific gravity of the urine becomes less according 
to the quantity of the sugar present. The urine is filtered if not 
clear. The specific gravity of the urine is determined by means 
of Sprengel's picnometer, which is provided with a thermometer 
and capillary tube. With this apparatus the specific gravity of 
the urine is determined with greater accuracy than with an 
ordinary picnometer or urinometer. 300 or 400 cc. of the urine, 
sp. gr. having been determined, is introduced into a flask, capacity 
1000 cc, and some yeast cake having been well washed with 
distilled water is mixed with the urine. Place the flask in 
a vessel containing water, a large water bath for example, so that 
it is surrounded by water, and keep the temperature of the 
water at 20 to 25 ° C. about 30 hours, with a microcosmic lamp. 
At the end of the process the yeast cells will have settled and the 
fluid be nearly clear. Filter through a dry filter paper into a dry 
flask, and bring the filtrate to the temperature at which the urine 
was when the specific gravity was determined, and ascertain its 
specific gravity with Sprengel's picnometer, taking care that the 
temperature be the same in both determinations. 

CALCULATION. 

Multiply the difference between the specific gravity of the 
urine before and after fermentation by 230, the product of which 
is the per cent, of sugar in the urine. If, however, the per cent, 
is less than 0.5, multiply by 219 instead of 230. 



THE OPTICAL METHOD. 

In almost all saccharimeters there are two Nicol's prisms, one . 
of which polarizes the light, and the other acts as an analyzer. 
A ray of light entering the polarizer is divided in two parts 
vibrating at right angles to each other, but the construction of 
the prism is such that the ray vibrating in one plane is absorbed 
while the other passes through. Light, therefore, which passes 
through and vibrates in one plane is polarized. Now, if the second 
prism or analyzer is related to the first so that their oblique 
ends are parallel, the polarized ray will pass through it without 



176 



CHEMICAL ANALYSIS OF THE URINE. 



obstruction ; but if rotated on its axis, the light passing through 
gradually becomes less until the rotation reaches 90 , when the 
light is cut off. From 90 , if the rotation be continued, light 
begins to pass, and the quantity increases until 180 is reached, 
when the polarized light again passes without obstruction, and by 
continuing the rotation from 180 , the light passing becomes less 
bright until the rotation is 270 , when no more light passes, and 
from this degree the light appears as the rotation takes place and 
passes without obstruction when the rotation is 360 , or the 



Fig. 36. 




original position of ihe prism. If the prisms are so related that the 
polarized light passes without obstruction, and there be placed 
between them a solution of grape or diabetic sugar in a glass tube, 
having its ends closed by glass plates, the plane of vibration of the 
light is rotated by the sugar so that when it falls on the second 
prism it is partly obstructed in its course ; however, by turning the 
latter on its axis, a degree of rotation is reached when the light 
passes through it. The plane of polarization is turned^ according 



THE OPTICAL METHOD. 177 

to the amount of sugar in solution, and the length of the tube con- 
taining the solution, but the difficulty of determining a.t what degree 
of rotation the greatest amount of light is transmitted is very 
great; hence, in nearly all saccharimeters means are employed by 
which it is at once determined when the light reaches the eye in 
the greatest quantity, one of the prisms being in connection with 
the arc of the circle or disc, which is graduated in degrees and 
minutes, so that when the prism is turned, either the disc or 
vernier is likewise turned. It is not necessary to enter here into a 
consideration of the devices employed in the various saccharime- 
ters in use.* For exact results, the apparatus of either Wild or 
Laurent is preferable, as the use of either does not depend on the 
ability to note differences in tints of color ; hence errors cannot 
arise from color blindness. In either apparatus the sodium light 



is employed. It is produced by sodium chloride, supported by a 
loop of platinum wire in a Bunsen's flame. Laurent, however, 
has constructed a special lamp — Laurent's Eolipile — for the 
production of the sodium light. The general arrangement of 
Wild's saccharimeter is represented by Fig. 36. In this apparatus 
the relationship of the two Nicol's prisms is known by the disap- 
pearance of black horizontal lines in the centre of the field of 
vision, as seen by b, Fig. 37. Crossing at acute angles are two 
lines, which remain distinctly in view at all times if the telescopic 
part of the polariscope is adjusted, which is near the eye piece. 
The graduated disc and the Nicol's prism by which the light is 
polarized are turned by means of the rod, C, the distal end of 
which is geared with the disc. The markings of the disc in 

* For an exhaustive treatment of the subject, the student is referred to Dr. Landolt's 
Handbook of the Polariscope and its Practical Applications, translated. MacMillan 
& Co. 



178 CHEMICAL ANALYSIS OF THE URINE. 

degrees are read by means, of the telescope, B, and they are 
made visible by a special lamp, D. Before estimating sugar in 
urine, the telescope is adjusted so the lines on the scale are well 
defined. This is accomplished by moving the ocular of the 
telescope forward or backward until the markings are plainly 
visible. The sodium flame having been placed in the axis of the 
saccharimeter, and one of the tubes, empty or filled with distilled 
water, placed in position, the telescopic part of the polariscope, A, is 
adjusted so the cross lines are distinctly seen when the graduated 
disc is turned by the rod, C, until the horizontal lines disappear 
from the centre of the field 6, Fig. 37. The instrument is 
adjusted if the vernier is found at o, 90, 180 or 270 . In 
determining when the horizontal lines disappear, the cross lines 
serve the purpose of ascertaining if the telescopic part of the 
polariscope, A, has remained in adjustment, or if the eye has 
changed in its accommodation ; in either case the telescopic part is 
adjusted so the cross lines are sharply defined, and if necessary 
the graduated disc is again turned until the horizontal lines 
disappear from the centre of the field of vision If»a variation is 
met, that is, if the vernier is not at o when the horizontal lines 
disappear from the central portion of the field of view, tests are 
made in the different quadrants of the circle, and in making 
estimations the variation is taken into account. 

In Laurent's apparatus the blue and violet rays of the sodium 
light are separated by the light passing through a plate of 
potassium bichromate. Half way across the path of the light, 
having been polarized, is a quartz plate of uniform and definite 
thickness. It is by means of this plate that, when the prisms 
sustain certain relationships, the half of the field of view crossed 
by the quartz plate is dark, while the other half is light, and by 
other relationships of the prisms both halves of the field of view 
are equally light. To adjust the instrument, the small cog wheel 
geared with the large graduated disc is turned to the .right or 
left until the field of vision becomes equally light, and the margin 
of the quartz plate disappears, as shown by a, Fig. 38. If the 
o of the vernier corresponds to that of the graduated disc, the 
instrument is adjusted. 

To estimate sugar in the urine it should be clear, and if not 
clear, it is filtered. Sugar in highly-colored urine cannot well be 



THE OPTICAL METHOD. 179 

estimated with a saccharimeter, and especially is this true with 
Laurent's apparatus. However, diabetic urine is generally free 
of much coloring matter, except in case of fever. To remove 
excess of coloring matter, shake the urine in a flask with a small 
quantity of pulverized neutral lead acetate and filter. To avoid 
diluting the urine, the flask should be dry and the filter not 
moistened with water before filtering. In filtering or decolorizing 
•urine, sufficient quantity is taken to examine with tubes of different 
lengths. Before filling the tubes, the water in them is removed by 
rinsing with the urine several times. The presence of water not only 
dilutes the urine, but, unless it is intimately mixed with the urine, 
it interferes with the passage of light. The effect which albumen 
has in changing the plane of vibration of polarized light is so 
inconsiderable that, when in small quantity in the urine, its 
removal is not necessary, but if in considerable quantity it should 
be removed; consequently, tests for albumen are made before 

Fig. 38. 



oca' 



estimating sugar with a saccharimeter. To separate albumen, 9 
volumes of urine (Hofmeister) are mixed with 1 volume of a solu- 
tion of sodium phosphortungstate which has been rendered strongly 
acid with hydrochloric acid. Having shaken the mixture in a 
bottle or flask, filter through a dry filter into a dry flask ; the 
tube, 10 cm. long, having been filled with urine and closed, to the 
exclusion of air, is placed in position, the sodium light having been 
brought in the axis of the saccharimeter and the vernier at o. In 
Wild's apparatus the graduated disc is turned to the left (in 
opposite direction to that of the hands of a clock) until the 
horizontal lines disappear from the centre of the field of view. As 
in adjusting the apparatus, attention is directed to the cross 
lines, and if they are indistinct, the telescopic part of the polari- 
scope, A, is adjusted before proceeding. In Laurent's apparatus the 
vernier is moved from above to the right, as the hands of a clock 



180 CHEMICAL ANALYSIS OF THE URINE. 

(this motion is imparted by turning the cog wheel to the left), 
to reach the degree at which the light of each half of the field of 
vision becomes equally bright, and the perpendicular margin of 
the quartz plate disappears from view. As a control of the result 
reached by use of the tube 10 cm. long with either apparatus, 
the tube 20 cm. long is employed, and the number of degrees 
rotated should be twice that of the first ; or dilute 1 vol. urine 
with 1 vol. water, and having mixed well examine with the tube 
10 cm. long; the number of degrees rotated should be one-half 
that with the urine before dilution. 

The calculation of the per cent, of sugar in urine from the 
number of degrees rotated with the tube 10 cm. long is quite 
simple. Multiply the number of degrees of rotation by 100 and 
divide by 53.1 (specific rotation of diabetic sugar), the quotient 
is the per cent, of sugar in the urine. 

-REMARKS. 

As normal urine contains substances which reduce copper 
oxide, the results of Fehling's method, or Pavy's modification of 
Fehling's method, are 0.2 to 0.4 per cent, too high. Roberts' 
method yields good results, and in many respects is prefer- 
able in estimating sugar in urine to that of Fehling or Pavy. 
It has the advantage of not requiring much apparatus. Instead 
of employing Sprengel's picnometer to determine the specific 
gravity, a urinometer may be used, when the results will be 
approximative. The results obtained by the Optical Method are 
the most accurate. If, in estimating sugar in the urine by 
Roberts' method and also by the Optical Method, the results are 
considerably higher by the former than by the latter, there 
is reason to believe that fruit sugar is present. Although fruit 
sugar has the same molecular constitution as diabetic sugar, and 
it undergoes fermentation and reduces CuO, yet, when in solu- 
tion, it changes the plane of vibration of polarized light to the 
left instead of the right ; as diabetic sugar, therefore, the results of 
the optical method are too low. In diabetic urine, however, this 
form of sugar is seldom found. 



APPENDIX. 



TABLE 1.— NITROGENOUS COMPOUNDS, EXCEPT UROBILIN, IN NORMAL 

URINE. 



Constituent. 



Number of gram- 
Average number mes nitrogen ex- 
p t fiv-r- Relative number of grammes of creted in 24 hours, 
tercen.ot. 1 10- of parts f eac ] 1 eac ^ const ituent calculated from 
constituent. excreted in 24 the per cent, of 
hours by adult of nitrogen in each 
average wt., etc. compound. 



Urea 



Uric Acid . . 
Ammonia . . . 
Kreatinin . . . 
Hippuric Acid . 
Indican. . . . 
Xanthin . . . 



46.66 
33-33 

82.35 

37.26 

7.87 

5-57 

36.85 



1.0 


31.0 


14.464 


001938 


0.6 


0.199 


0.02257 


0.7 1 


0.576 


02257 


0.7 


0.260 


0.01129 


0.4 


0.031 


0.00035 


O.OII 


0.0006 


— 


0.0056 


0.00206 



Total number grms. nitrogen excreted in 24 hours by adult of 

average weight, etc 15.53266 

Average number of grms. nitrogen of urea excreted in 24 hours by 

adult of average weight, etc 14.4640 

Average number of grms. nitrogen of compounds other than urea 

excreted in 24 hours by adult of average weight 1.06866 

Per cent, of nitrogen in combination in urea of the total quantity 

excreted 93- 12 

Per cent, of nitrogen in combination in other compounds than urea 

of the total quantity (100 per cent.) excreted 6.88 

181 



182 



APPENDIX. 



TABLE 2.— (.FROM ZUELZER'S "SEMIOLOGIE DES HARNS.")— NUMBER OF PARTS 
OF NITROGEN AND OTHER CONSTITUENTS OF ELEMENTS OF FOOD IN 
iooo PARTS. 



Element of Food. Nitrogen. P 2 O s . H 2 S0 4 K. 



Muscular tissue 
of the ox. . . 

Calf 

Horse .... 

Brain (all parts 
of) of the ox. 



34.6 4.45 , 5.6 ! 3:6 



Na. 



O.58 



MgO. CaO. 



CI. 



O.29 O.16 O.69] 



157 



3-73 



2.19 0.43 0.15 0.13 0.5 



50.19 4.85 8.2 



2.8 



0.71 



0.107 0.202 



7.416 trace 3.26 1.32 0.135 0.36 0.514 



Milk (human) . 


5-4 


P. 43 7 


0.07 


0.7 


0.09 


0.02 


0.431 0.437 


Milk (cow) . . 


3.8i 


2.1 


°-395 


0.875 


0.47 


0.299 


1.864 o.75 


Wheat 


20.8 


7-9 


0.12 


4-3 


0.29 


2.00 


0.60 0.03 


Oats 


19.2 


6.2 


0.49 


3-6 


0.44 


i-9 


1. 00 0.15 


Potatoes . . . 


3-2 


1.6 


0.S5 


4.8 


0.12 


0.4 


0.20 0.29 



TABLE 3.— (FROM ZUELZER'S " SEMIOLOGIE DES HARNS.")-RELATIVE NUMBER 
OF PARTS OF CONSTITUENTS OF CERTAIN ANIMAL AND VEGETABLE 
BODIES FOR 100 PARTS OF NITROGEN WHICH THEY CONTAIN. 





p 2 o 5 . 


H 2 S0 4 


K. 


Na. 


MgO. 


CaO. 


CI. 


Muscular tissue (human). . 


12. 1 


23.1 


9.0 


4-5 


I.I 


0.6 





Muscular tissue (horse). . 


15-7 


24.O 


9.1 


2-3 


1.4 


0.4 


o-5 


Muscular tissue (ox) . . . 


12.8 


16.7 


10.4 


1.6 


i-5 


0.6 


1.9 


Muscular tissue (calf). . . 


10.4 





6.1 


1.1 


0.4 


o-3 


1.4 


Brain (average) 


44.0 


0.7 


21.0 


8.7 


1.1 


0.6 


2.6 


Blood from all parts of body. 





i-5 


3-5 


6.0 


1.0 


3-o 


6.0 


Milk (human) 


13-4 


2.0 


18.8 


4.4 


1.4 


10.7 


12.9 


Milk (cow) 


55-i 


10.3 


23.4 


12.3 


7-9 


49.0 


19.7 


Wheat 


38.0 


0.57 


20.6 


I -3 


9.6 


2.8 


O.I 


Oats 


37-5 


2.5 


18.7 


2.2 


9-8 


5-2 


0.7 



APPENDIX. 



183 



TABLE 4.— (FROM ZUELZER'S "SEMIOLOGIE DES HARNS.")— INDICATING 
CHANGES IN THE CONSTITUTION OF THE URINE BY THE INGESTION 
OF DIFFERENT KINDS OF FOOD. NUMBER OF PARTS FOR 106 PARTS 
OF NITROGEN EXCRETED. 





p 2 o B . 


H 3 S0 4 . 


MgO. 


CaO. 


K 


Na. 


Blood . . . 


ii. 9 


19.0 


I.O 


0.4 


16.6 


II. 2 


Beef .... 


II.O 


17. 1 


O.4 


0.2 


9.6 


4.I 


Horse flesh . 


15-4 


16.8 


0-5 


0.7 






Kidney . . 


23.0 


25.6 


0.5 


0.4 








Liver, cooked 


26.3 


18.8 


0.4 


0.5 


IO.8 


IO.8 


Liver, fresh . 


3*-3 


24.9 





0.4 


12.4 


6.5 


Brain . . . 


S3-* 


10.8 


O.08 


0.4 







Brain . . . 


30.8 


25-5 


O.7 


0.8 


22.7 


88 


Milk .... 


21.3 


ro. 


0.8 


1.2 


34-i 


33 9 



INDEX. 



Acid Urine, Sediments peculiar to... 70 

Acidity of Urine, Estimation of 118 

Albumen 42 

Tests for 46 

Removal of, in testing for sugar 55 

in Process of Analysis 86 

Estimation of, Scherers Method 166 

Albuminous Bodies 42 

Color Tests for 46 

Estimation of 166 

Alkaline Urine, Sediments peculiar to 72 
see Properties of Urine, 
see Fermentation of Urine. 

Ammonia , 39 

Urate of, in Sediments 74 

" " Concretions 95 

see Table 1, Appendix. 
Estimation of, (Schlosing and 
Xeubauer) 159 

Analysis of Concretions 96 

Sediments 91 

Urine 85 

Quantitative 101 

Bacteria 82 

Balance of Precision 104 

Barium Mixture, Preparation of 115 

Benzoic Acid 29 

Origin of 29 

Biliary Acids, Properties of 57 

Pettenkoffer's Test for 57 

Separation of, from Urine 58 

in Process of Analysis 87 

Coloring Matters 58 

Gmelin's Test for.. 59 

in Process of Analysis 87 

Bilirubin, see Biliary Coloring Matters. 

Blood 63 

Corpuscles, 80 

Coloring Matters of. 60 

Spectroscopic Test for 60, 61 
see Hrematin. 

Burettes t 106 



Calcium Phosphate Basic Sediments 73 



Oxalate 



in Concretions 95 

Oxide, Estimation of, Gravimetric 

Method 161 

Oxide, Estimation of, Volumetric 

Method 162 

Casts, Epithelial, in Sediments 77 

" " Process of Analysis 93 

Hyaline, ' ; Sediments 77 

" " Process of Analysis 93 

Waxy, " Sediments 78 

" " Process of Analysis 93 

Blood, " Sediments 79 

" " Process of Analysis 93 

Chlorine 32 

in Process of Analysis 88 

Estimation of, Volhard and Sal- 

kowski 153 

Estimation of, Xeubauer 156 

Cholesterin 67 

Separation of, from Urine 67 

Color Tests for Albuminous Bodies 44, 46 

Coloring Matters of Urine 27 

Estimation of 118 

Concretions 95 

Scheme for the Analysis of. 96 

Crucible, Platinum 10S 

Porcelain 10S 

Cystin 76 

in Concretions 95 

Desiccators 108 

Diabetes Mellitus 49 

Dish, Platinum 10S 

Drying 103 

Dumas' Method, see Xitrogen. 

Epithelial Cells 79 

Casts 77 

Ester Compounds of Sulphur 36 

Evaporating 102 

Streng's Method of 63 



Calcium 38 Fat 66 

Sulphate, in Sediments 72 in Process of Analysis 86 

Phosphate Xeutral " 72 Fehling's Solution 50 

185 



186 



INDEX. 



PAGE 

Fermentation of Urine 20 

Test for Sugar 52 

in Estimation of Sugar, Roberts' 

Method 175 

Fibrin 64 

Filter Paper 101 

Filtering 101 

Filter Syphon 150 

Globuline. 45 

in Process of Analysis 87 

Estimation of, Hammarsten's 

Method 167 

Estimation of, Pohl's Method.. 168 

Glycerin-Phosphoric Acid 34 

Test for, (Sotnischewsky) 34 

from Lecithin 67 

in Process of Analysis 88 

Estimation of 146 

Gonorrhoea, Sediments in 82 

Gravel, see Concretions. 

Guanin. see Xanthin Group. 

Hematin, Heller's Test for : 63 

Struve's " " 63 

Hemoglobin 60 

H emialbu mose 45 

Color Tests for 44, 46 

in Process of Analysis 87 

Estimation of 169 

Hellers Test, see Haematin. 

Hippuric Acid 25 

Origin of.. 25 

in Sediments 71 

" Process of Analysis 91 

Hydrochloric Acid, Normal 113 

Indican 26 

Test for (Jaffe) 27 

Indoxylsulphuric Acid, see 
Indican. 

Inorganic Bodies 31, 117 

in Concretions 95 

Estimation of.. 117 

Kreatinin 23 

Zinc Chloride 24 

see Table 1, Appendix. 
Estimation of (Neubauer and 

Salkowski) 139 

Kjeldahl's Method, see Nitrogen. 

Lecithin, Decomposition of, and Test 

for 67 

see Glycerin-Phosphoric Acid. 

Leucin 64 

Litmus Solution 114 

Magnesium 3& 

Phosphate, see Phosphoric Acid. 



PAGE 

Magnesium Ammon. Phosphate, in 

Sediments 72 

Ammon. Phosphate, in Process 

of Analysis 92 

Mixture 115 

in Concretions 95 

Oxide, Estimation of, Gravi- 
metric Method 63 

Oxide, Estimation of, Volumetric 

Method 164 

Methsemoglobin 60 

Micrococcus Ure?e. see Fermentation 
of Urine. 

in Process of Analysis 94 

Microorganisms 82 

Microscopic Examination of Sedi- 
ments 89 

Millon's Reagent, Preparation of 115 

Mould (Penicilium Glaucum) 84 

Neurin, Tests for 6S 

Nitric Acid Test for Albumen 46 

Nitrogen, Estimation of, in Urine... 128 
Nitrogenous Bodies in Urine, see 
Table 1, Appendix. 

Odor of Urine . see Properties of Urine. 
Optical Method, see Estimation of 
Sugar. 

see Estimation of Hemialbu- 

mose. 
see Estimation of Peptone. 

Organized Sediments 77 

Oxalic Acid 29 

Estimation of 141 

in Sediments, see Calcium 
Oxalate. 

in Concretions 95 

Normal 109 

Oxyhemoglobin 60 

Pavy's Solution 174 

Peptone 43, 45 , 46 

ColorTests for 44, 46 

in Process of Analysis 87 

Estimation of 169 

Phenylhydrazin Test for Sugar 53 

Phosphates, Constitution of 32 

Deportment of 33, 34 

in Sediments 72, 73 

in Process of Analysis 88, 92 

in Concretions 95 

Phosphoric Acid 32 

Estimation of 143 

see Glycerin- Phosphoric Acid. 

Phosphorus Compounds 32 

in Urine by Ingestion of Foods, 
see Table 4, Appendix. 

Pipettes 106 



INDEX. 



18* 



PAGE 

Potassium 4° 

Estimation of 157 

Hydrate, Normal 109 

Properties of Urine 17 

Pus 80 

Qualitative Analysis of Urine, 

Scheme for 85 

of Sediments, Scheme for 91 

of Concretions, " " 96 

Quantitative Analysis 85 

Reactions of Urine, see Properties 

of Urine. 
Roberts' Method, see Estimation of 

Sugar. 

Saccharimeters, Wild's and Laurent's 

176, 178 

Sarcina 83 

Sarkin. see Xanthin Group. 

Sediments in Urine 70 

Silver Nitrate, Standardized Solution 

of 154 

Specific Gravity of Urine 18, 116 

Spectroscope 61 

Spermatozoa 82 

Staining Fluids 90 

Stones, see Concretions. 

Sugar, Diabetic 49 

Trommer and Salkowski's Test 

for 50 

Test for with Fehling's Solution 50 

" by Fermentation 52 

" with Phenylhydrazin... 53 

Separation of, from Urine 54 

in Albuminous Urine 55 

in Process of Analysis 87 

Estimation of Fehling's Method 170 
" " and Pavy's 

Method 173 

Estimation of Roberts' Method 175 
" Optical " 175 
Remarks on Methods of Esti- 
mating 180 

Sulphur Compounds in Urine 36 

Estimation of Total Quantity of 

147, 152 

Sulphuric Acid as Sulphates 36 

as Esters 36 

Estimation of, Gravimetric Me- 
thod 147 

Estimation of, Volumetric Me- 
thod 149 

Preparation of Normal...-. 112 



I'AGE 

Table Illustrating Spectra of Coloring 

Matters of Blood, etc 60 

of Specific Gravity, of Dilute 

KOH solutions no 

of Specific Gravity, of Dilute 

NaOH solutions in 

of Specific Gravity, of Dilute 

H 2 S0 4 solutions ; 113. 

of Specific Gravity, of Dilute 

HC1 solutions 114 

Tyrosin 65 

Test for 65 

in Sediments 77 

in Process of Analysis 88- 

Urates 23 

in Sediments 70 

" Process of Analysis 88,91 

" Concretions. 95 

Urea 19 

in Fermentation of Urine 21 

Consult Table 1, Appendix. 
Estimation of, Liebig and Pflii- 

ger's Method 119 

Estimation of, in Diseased 

Urine 125 

Estimation of, Knop and 

Greene's Method 125 

Uric Acid 22 

in Sediments 70 

Test for (Murexide Test) 23 

in Process of Analysis 91 

Consult Table I, Appendix. 

in Concretions 95. 

Estimation of, Salkowski and 

Ludwig's Method 138 

Urobilin 27 

Varrentrapp and Will's Method. 

see Nitrogen. 
Volume of Urine Excreted in 24 

hours 116 

Volumetric Analysis, Preparation 

of Solutions for 108 

Weights 104 

Weyl's Test for Kreatinin 25 

Wild's Saccharimeter 176 

Xanthin 21 

in Concretions 95. 

Consult Table I, Appendix. 

Group 21 

Zulkowsky's Azotometer 130 



